CORD-19-Sentences  

CORD-19:acb5f1a150a07ca259285e3558c2c9a1869fd6af JSONTXT 9 Projects

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Id Subject Object Predicate Lexical cue
TextSentencer_T1 0-76 Sentence denotes Characterization of leader-related small RNAs in coronavirus-infected cells:
TextSentencer_T1 0-76 Sentence denotes Characterization of leader-related small RNAs in coronavirus-infected cells:
TextSentencer_T2 77-138 Sentence denotes Further evidence for leader-primed mechanism of transcription
TextSentencer_T2 77-138 Sentence denotes Further evidence for leader-primed mechanism of transcription
TextSentencer_T3 140-148 Sentence denotes Abstract
TextSentencer_T3 140-148 Sentence denotes Abstract
TextSentencer_T4 149-340 Sentence denotes Mouse hepatitis virus (MHV), a murine coronavirus, replicates in the cytoplasm and synthesizes 7 viral mRNAs containing an identical stretch of leader RNA sequences at the 5'-end of each RNA.
TextSentencer_T4 149-340 Sentence denotes Mouse hepatitis virus (MHV), a murine coronavirus, replicates in the cytoplasm and synthesizes 7 viral mRNAs containing an identical stretch of leader RNA sequences at the 5'-end of each RNA.
TextSentencer_T5 341-492 Sentence denotes The leader-coding sequences at the 5'-end of genomic RNA are at least 72 nucleotides in length and are joined to the viral mRNAs by a unique mechanism.
TextSentencer_T5 341-492 Sentence denotes The leader-coding sequences at the 5'-end of genomic RNA are at least 72 nucleotides in length and are joined to the viral mRNAs by a unique mechanism.
TextSentencer_T6 493-626 Sentence denotes Utilizing a leader-specific cDNA probe, we have detected several free leader RNA species ranging from 70 to 82 nucleotides in length.
TextSentencer_T6 493-626 Sentence denotes Utilizing a leader-specific cDNA probe, we have detected several free leader RNA species ranging from 70 to 82 nucleotides in length.
TextSentencer_T7 627-687 Sentence denotes The predominant leader RNA was approximately 75 nucleotides.
TextSentencer_T7 627-687 Sentence denotes The predominant leader RNA was approximately 75 nucleotides.
TextSentencer_T8 688-805 Sentence denotes In addition, larger distinct leader-containing RNAs were also detected ranging from 130 to 250 nucleotides in length.
TextSentencer_T8 688-805 Sentence denotes In addition, larger distinct leader-containing RNAs were also detected ranging from 130 to 250 nucleotides in length.
TextSentencer_T9 806-921 Sentence denotes The 70-82-nucleotide leader-related RNAs were present in both the cytosol and membrane fractions of infected cells.
TextSentencer_T9 806-921 Sentence denotes The 70-82-nucleotide leader-related RNAs were present in both the cytosol and membrane fractions of infected cells.
TextSentencer_T10 922-1035 Sentence denotes They were also detected only in the small RNA fractions but not associated with the replicative-intermediate RNA.
TextSentencer_T10 922-1035 Sentence denotes They were also detected only in the small RNA fractions but not associated with the replicative-intermediate RNA.
TextSentencer_T11 1036-1203 Sentence denotes These data suggest that the leader RNAs were associated with the membrane-bound transcription complex but at least part of them were dissociated from the RNA template.
TextSentencer_T11 1036-1203 Sentence denotes These data suggest that the leader RNAs were associated with the membrane-bound transcription complex but at least part of them were dissociated from the RNA template.
TextSentencer_T12 1204-1419 Sentence denotes We have also identified a temperature-sensitive mutant, which synthesizes only leader RNA but not mRNAs at nonpermissive temperature, indicating that leader RNA synthesis is distinct from the transcription of mRNAs.
TextSentencer_T12 1204-1419 Sentence denotes We have also identified a temperature-sensitive mutant, which synthesizes only leader RNA but not mRNAs at nonpermissive temperature, indicating that leader RNA synthesis is distinct from the transcription of mRNAs.
TextSentencer_T13 1420-1581 Sentence denotes These data support the leader-primed mechanism for coronavirus transcription and suggest that one or more free leader RNAs are used as primers of mRNA synthesis.
TextSentencer_T13 1420-1581 Sentence denotes These data support the leader-primed mechanism for coronavirus transcription and suggest that one or more free leader RNAs are used as primers of mRNA synthesis.
TextSentencer_T14 1582-1770 Sentence denotes Mouse hepatitis virus, transcription, small leader related RNAs, coronavirus, temperature-sensitive mutant 0168-1702/85/$03.30 0 1985 Elsevier Science Publishers B.V. (Biomedical Division)
TextSentencer_T14 1582-1770 Sentence denotes Mouse hepatitis virus, transcription, small leader related RNAs, coronavirus, temperature-sensitive mutant 0168-1702/85/$03.30 0 1985 Elsevier Science Publishers B.V. (Biomedical Division)
TextSentencer_T15 1772-1926 Sentence denotes Mouse hepatitis virus (MHV), the prototype of Coronaviridae, contains a singlestranded and plus-sensed RNA of 5.4 X lo6 daltons (Lai and Stohlman, 1978) .
TextSentencer_T15 1772-1926 Sentence denotes Mouse hepatitis virus (MHV), the prototype of Coronaviridae, contains a singlestranded and plus-sensed RNA of 5.4 X lo6 daltons (Lai and Stohlman, 1978) .
TextSentencer_T16 1927-2081 Sentence denotes The 60s genome is enclosed within a helical nucleocapsid constructed from multiple copies of capsid protein, pp60 (Sturman, 1977; Stohlman et al., 1983) .
TextSentencer_T16 1927-2081 Sentence denotes The 60s genome is enclosed within a helical nucleocapsid constructed from multiple copies of capsid protein, pp60 (Sturman, 1977; Stohlman et al., 1983) .
TextSentencer_T17 2082-2315 Sentence denotes During maturation, the nucleocapsid buds into endoplasmic reticulum, forming an intact virion containing an envelope and two virus-specific glycoproteins, gp 23 and gp 180/90 (Sturman, 1977; Wege et al., 1979; Siddell et al., 1981) .
TextSentencer_T17 2082-2315 Sentence denotes During maturation, the nucleocapsid buds into endoplasmic reticulum, forming an intact virion containing an envelope and two virus-specific glycoproteins, gp 23 and gp 180/90 (Sturman, 1977; Wege et al., 1979; Siddell et al., 1981) .
TextSentencer_T18 2316-2461 Sentence denotes Murine coronaviruses are normally associated with hepatitis in vivo and infect a variety of established cell lines in vitro (Wege et al., 1982) .
TextSentencer_T18 2316-2461 Sentence denotes Murine coronaviruses are normally associated with hepatitis in vivo and infect a variety of established cell lines in vitro (Wege et al., 1982) .
TextSentencer_T19 2462-2590 Sentence denotes These viruses replicate in the cytoplasm, independently of nuclear involvement (Brayton et al., 1981; Wilhelmsen et al., 1981) .
TextSentencer_T19 2462-2590 Sentence denotes These viruses replicate in the cytoplasm, independently of nuclear involvement (Brayton et al., 1981; Wilhelmsen et al., 1981) .
TextSentencer_T20 2591-2762 Sentence denotes Upon entry, the viral genome is translated into an 'early' polymerase which directs the synthesis of full-length (-)-sensed RNA (Lai et al., 1982b; Brayton et al., 1984) .
TextSentencer_T20 2591-2762 Sentence denotes Upon entry, the viral genome is translated into an 'early' polymerase which directs the synthesis of full-length (-)-sensed RNA (Lai et al., 1982b; Brayton et al., 1984) .
TextSentencer_T21 2763-2918 Sentence denotes This enzymatic activity is replaced later in infection by two separate 'late' polymerase activities which synthesize either genomic or mRNAs, respectively.
TextSentencer_T21 2763-2918 Sentence denotes This enzymatic activity is replaced later in infection by two separate 'late' polymerase activities which synthesize either genomic or mRNAs, respectively.
TextSentencer_T22 2919-3032 Sentence denotes These enzymatic activities are associated with cellular membranes (Brayton et al., 1982 (Brayton et al., , 1984 .
TextSentencer_T22 2919-3032 Sentence denotes These enzymatic activities are associated with cellular membranes (Brayton et al., 1982 (Brayton et al., , 1984 .
TextSentencer_T23 3033-3232 Sentence denotes Seven virus-specific mRNAs ranging from 0.6 to 5.4 X 10' daltons are detected on polysomes of virus-infected cells (Lai et al., 1981 (Lai et al., , 1982a Leibowitz et al., 1981; Spaan et al., 1981) .
TextSentencer_T23 3033-3232 Sentence denotes Seven virus-specific mRNAs ranging from 0.6 to 5.4 X 10' daltons are detected on polysomes of virus-infected cells (Lai et al., 1981 (Lai et al., , 1982a Leibowitz et al., 1981; Spaan et al., 1981) .
TextSentencer_T24 3233-3448 Sentence denotes These RNAs are arranged in the form of a nested set from the 3'-end of the genome such that the sequences of each smaller RNA are contained within the 3'-sequences of the next larger RNA species (Lai et al., 1981) .
TextSentencer_T24 3233-3448 Sentence denotes These RNAs are arranged in the form of a nested set from the 3'-end of the genome such that the sequences of each smaller RNA are contained within the 3'-sequences of the next larger RNA species (Lai et al., 1981) .
TextSentencer_T25 3449-3699 Sentence denotes In addition to this unique genetic arrangement, it has recently been demonstrated that each mRNA and genomic RNA contain an identical stretch of approximately 72 nucleotides at their 5'-ends (Lai et al., 1983 (Lai et al., , 1984 Spaan et al., 1983) .
TextSentencer_T25 3449-3699 Sentence denotes In addition to this unique genetic arrangement, it has recently been demonstrated that each mRNA and genomic RNA contain an identical stretch of approximately 72 nucleotides at their 5'-ends (Lai et al., 1983 (Lai et al., , 1984 Spaan et al., 1983) .
TextSentencer_T26 3700-3970 Sentence denotes The presence of a leader RNA common to all viral mRNAs indicates that they are synthesized by joining two noncontiguous RNA segments, since the leader sequences are encoded only at the 5'-end of the genomic RNA (Lai et al., 1983 (Lai et al., , 1984 Spaan et al., 1983) .
TextSentencer_T26 3700-3970 Sentence denotes The presence of a leader RNA common to all viral mRNAs indicates that they are synthesized by joining two noncontiguous RNA segments, since the leader sequences are encoded only at the 5'-end of the genomic RNA (Lai et al., 1983 (Lai et al., , 1984 Spaan et al., 1983) .
TextSentencer_T27 3971-4059 Sentence denotes These data suggest that a unique mechanism of RNA synthesis occurs during MHV infection.
TextSentencer_T27 3971-4059 Sentence denotes These data suggest that a unique mechanism of RNA synthesis occurs during MHV infection.
TextSentencer_T28 4060-4269 Sentence denotes This mechanism differs from conventional eukaryotic RNA splicing since UV transcriptional mapping studies indicate that these mRNAs are not derived from cleavage of large precursor RNAs (Jacobs et al., 1981) .
TextSentencer_T28 4060-4269 Sentence denotes This mechanism differs from conventional eukaryotic RNA splicing since UV transcriptional mapping studies indicate that these mRNAs are not derived from cleavage of large precursor RNAs (Jacobs et al., 1981) .
TextSentencer_T29 4270-4447 Sentence denotes The study of replicative-intermediate RNA further suggests that the leader RNA is joined to the mRNAs during transcription, but not post-transcriptionally (Baric et al., 1983) .
TextSentencer_T29 4270-4447 Sentence denotes The study of replicative-intermediate RNA further suggests that the leader RNA is joined to the mRNAs during transcription, but not post-transcriptionally (Baric et al., 1983) .
TextSentencer_T30 4448-4743 Sentence denotes We have also shown that the double-stranded replicative form (RF) is of genome length, suggesting that the joining mechanism does not involve 'looping out' of intervening sequences in the (-)-stranded RNA template since such a mechanism would have generated subgenomic RFs (Baric et al., 1983) .
TextSentencer_T30 4448-4743 Sentence denotes We have also shown that the double-stranded replicative form (RF) is of genome length, suggesting that the joining mechanism does not involve 'looping out' of intervening sequences in the (-)-stranded RNA template since such a mechanism would have generated subgenomic RFs (Baric et al., 1983) .
TextSentencer_T31 4744-5019 Sentence denotes Thus, these data are compatible with a model in which the coronavirus leader RNA is synthesized independently and then sequestered to prime transcription of the body sequences of each mRNA by binding specific initiation sites on the (-)-strand template (Baric et al., 1983) .
TextSentencer_T31 4744-5019 Sentence denotes Thus, these data are compatible with a model in which the coronavirus leader RNA is synthesized independently and then sequestered to prime transcription of the body sequences of each mRNA by binding specific initiation sites on the (-)-strand template (Baric et al., 1983) .
TextSentencer_T32 5020-5312 Sentence denotes This unique mechanism is supported by sequence data from mRNAs 6 and 7 (Armstrong et al., 1984; Spaan et al., 1983; Skinner and Siddell, 1981) , which demonstrate the presence of sequence complementarity between the 3'-portion of the leader RNA and intergenic 'start' sites on the (-) strand.
TextSentencer_T32 5020-5312 Sentence denotes This unique mechanism is supported by sequence data from mRNAs 6 and 7 (Armstrong et al., 1984; Spaan et al., 1983; Skinner and Siddell, 1981) , which demonstrate the presence of sequence complementarity between the 3'-portion of the leader RNA and intergenic 'start' sites on the (-) strand.
TextSentencer_T33 5313-5558 Sentence denotes Presently, the exact length of the leader RNA is not known but must consist of 57 nucleotides from the S-end of the genome plus at most 15 nucleotides which are complementary to the splice junctions on the (-)-stranded RNA (Spaan et al., 1983) .
TextSentencer_T33 5313-5558 Sentence denotes Presently, the exact length of the leader RNA is not known but must consist of 57 nucleotides from the S-end of the genome plus at most 15 nucleotides which are complementary to the splice junctions on the (-)-stranded RNA (Spaan et al., 1983) .
TextSentencer_T34 5559-5686 Sentence denotes In this communization, we demonstrate the synthesis and characterization of small leader-containing RNAs in M~V-infected cells.
TextSentencer_T34 5559-5686 Sentence denotes In this communization, we demonstrate the synthesis and characterization of small leader-containing RNAs in M~V-infected cells.
TextSentencer_T35 5687-5851 Sentence denotes Moreover, we have isolated a temperature-sensitive mutant which accumulates a leader RNA of approximately 72 nucleotides in length at the nonpermissive temperature.
TextSentencer_T35 5687-5851 Sentence denotes Moreover, we have isolated a temperature-sensitive mutant which accumulates a leader RNA of approximately 72 nucleotides in length at the nonpermissive temperature.
TextSentencer_T36 5852-6018 Sentence denotes These data support the 'leader-primed' mechanism for coronavirus transcription (Baric et al., 1983) and demonstrate another mode of RNA synthesis in eukaryotic cells.
TextSentencer_T36 5852-6018 Sentence denotes These data support the 'leader-primed' mechanism for coronavirus transcription (Baric et al., 1983) and demonstrate another mode of RNA synthesis in eukaryotic cells.
TextSentencer_T37 6019-6072 Sentence denotes The implications of these findings will be discussed.
TextSentencer_T37 6019-6072 Sentence denotes The implications of these findings will be discussed.
TextSentencer_T38 6073-6173 Sentence denotes The A59 strain of MHV was prupagated in DBT or L2 cells as previously described (Lai et al., 1981) .
TextSentencer_T38 6073-6173 Sentence denotes The A59 strain of MHV was prupagated in DBT or L2 cells as previously described (Lai et al., 1981) .
TextSentencer_T39 6174-6295 Sentence denotes Temperature-sensitive mutants of A59 were isolated on L2 cells following mutagenesis with 5-fluorouracil or 5azacytidine.
TextSentencer_T39 6174-6295 Sentence denotes Temperature-sensitive mutants of A59 were isolated on L2 cells following mutagenesis with 5-fluorouracil or 5azacytidine.
TextSentencer_T40 6296-6413 Sentence denotes The detailed protocol on mutant isolation and description of phenotypes of these mutants will be published elsewhere.
TextSentencer_T40 6296-6413 Sentence denotes The detailed protocol on mutant isolation and description of phenotypes of these mutants will be published elsewhere.
TextSentencer_T41 6414-6523 Sentence denotes Experiments utilizing these mutants were performed at permissive (32°C) and nonpermissive (39%) temperatures.
TextSentencer_T41 6414-6523 Sentence denotes Experiments utilizing these mutants were performed at permissive (32°C) and nonpermissive (39%) temperatures.
TextSentencer_T42 6524-6656 Sentence denotes The intracellular RNA from A5Pinfected cells was isolated between 5 and 7 h post-infection in the presence of 2 pg/ml actinomycin D.
TextSentencer_T42 6524-6656 Sentence denotes The intracellular RNA from A5Pinfected cells was isolated between 5 and 7 h post-infection in the presence of 2 pg/ml actinomycin D.
TextSentencer_T43 6657-6752 Sentence denotes The RNA was extracted with phenol/SDS as described in earlier publications (Lai et al., 1981) .
TextSentencer_T43 6657-6752 Sentence denotes The RNA was extracted with phenol/SDS as described in earlier publications (Lai et al., 1981) .
TextSentencer_T44 6753-6936 Sentence denotes The synthetic oligodeoxyribonucleotide (15mer) (5'-AGTTTAGATTAGA'TT-3') was prepared by Dr. Bruce Kaplan, Department of Molecular Genetics, City of Hope Medical Center, Duarte, Calif.
TextSentencer_T44 6753-6936 Sentence denotes The synthetic oligodeoxyribonucleotide (15mer) (5'-AGTTTAGATTAGA'TT-3') was prepared by Dr. Bruce Kaplan, Department of Molecular Genetics, City of Hope Medical Center, Duarte, Calif.
TextSentencer_T45 6937-7124 Sentence denotes The 1 S-mer was 5'-end-labeled with [ y-32P]ATP by polynucleotide kinase and hybridized to total intracellular RNA from the ~HV-infected cells as previously described (Lai et al., 1984) .
TextSentencer_T45 6937-7124 Sentence denotes The 1 S-mer was 5'-end-labeled with [ y-32P]ATP by polynucleotide kinase and hybridized to total intracellular RNA from the ~HV-infected cells as previously described (Lai et al., 1984) .
TextSentencer_T46 7125-7367 Sentence denotes After ethanol precipitation and several washings, leader-specific cDNA was synthesized with reverse transcriptase in a buffer containing 1 mM each of dATP, dGTP, TTP and dCTP, 10 mM MgCl,, 120 mM KC1 and 30 mM mercaptoethanol at 42*C for 1 h.
TextSentencer_T46 7125-7367 Sentence denotes After ethanol precipitation and several washings, leader-specific cDNA was synthesized with reverse transcriptase in a buffer containing 1 mM each of dATP, dGTP, TTP and dCTP, 10 mM MgCl,, 120 mM KC1 and 30 mM mercaptoethanol at 42*C for 1 h.
TextSentencer_T47 7368-7548 Sentence denotes The RNA template was removed by alkaline treatment, and the full-length cDNA product (72 nucleotides long) was isolated by molecular sieve chromatography on a Sephadex G-50 column.
TextSentencer_T47 7368-7548 Sentence denotes The RNA template was removed by alkaline treatment, and the full-length cDNA product (72 nucleotides long) was isolated by molecular sieve chromatography on a Sephadex G-50 column.
TextSentencer_T48 7549-7687 Sentence denotes The leader-specific cDNA was synthesized as described above, except that 32 Ia-PI dCTP was used instead of unlabeled dCTP in the reaction.
TextSentencer_T48 7549-7687 Sentence denotes The leader-specific cDNA was synthesized as described above, except that 32 Ia-PI dCTP was used instead of unlabeled dCTP in the reaction.
TextSentencer_T49 7688-7957 Sentence denotes The products were separated on 12% polya~~la~de gels containing 6 M urea, located by autoradiogra-phy, and the 32P-labeled 72-nucleotide leader cDNA eluted in DNA extraction buffer (0.5 M ammonium acetate, 0.01 M Mg acetate, 0.001 M EDTA and 0.1% SDS) for 12 h at 65°C.
TextSentencer_T49 7688-7957 Sentence denotes The products were separated on 12% polya~~la~de gels containing 6 M urea, located by autoradiogra-phy, and the 32P-labeled 72-nucleotide leader cDNA eluted in DNA extraction buffer (0.5 M ammonium acetate, 0.01 M Mg acetate, 0.001 M EDTA and 0.1% SDS) for 12 h at 65°C.
TextSentencer_T50 7958-8176 Sentence denotes Following ethanol precipitation, the cDNA was hybridized to the 60s genomic RNA isolated from the purified virus in 80% formamide, 0.4 M NaCl, 0.01 M Na phosphate (pH 7.0) and 0.01 M Pipes-HCl (pH 6.8) at 37°C for 5 h.
TextSentencer_T50 7958-8176 Sentence denotes Following ethanol precipitation, the cDNA was hybridized to the 60s genomic RNA isolated from the purified virus in 80% formamide, 0.4 M NaCl, 0.01 M Na phosphate (pH 7.0) and 0.01 M Pipes-HCl (pH 6.8) at 37°C for 5 h.
TextSentencer_T51 8177-8353 Sentence denotes The hybrid was washed extensively and digested with 1000 units of S, nuclease in 0.2 M NaCl, 0.05 M Na acetate (pH 4.5) 0.0018 M zinc sulfate and 0.5% glycerol for 1 h at 37°C.
TextSentencer_T51 8177-8353 Sentence denotes The hybrid was washed extensively and digested with 1000 units of S, nuclease in 0.2 M NaCl, 0.05 M Na acetate (pH 4.5) 0.0018 M zinc sulfate and 0.5% glycerol for 1 h at 37°C.
TextSentencer_T52 8354-8492 Sentence denotes The reaction mixture was adjusted to 0.01 M EDTA and extracted with phenol/chloroform prior to electrophoresis on 12% polyacrylamide gels.
TextSentencer_T52 8354-8492 Sentence denotes The reaction mixture was adjusted to 0.01 M EDTA and extracted with phenol/chloroform prior to electrophoresis on 12% polyacrylamide gels.
TextSentencer_T53 8493-8595 Sentence denotes Infected cells, harvested between 5 and 6 h post-infection, were swollen in 0.5 X RSB buffer (1 X RSB:
TextSentencer_T53 8493-8595 Sentence denotes Infected cells, harvested between 5 and 6 h post-infection, were swollen in 0.5 X RSB buffer (1 X RSB:
TextSentencer_T54 8596-8647 Sentence denotes 0.01 M Tris-HCl, pH 7.4/0.01 M NaCl/0.0015 M MgCl,)
TextSentencer_T54 8596-8647 Sentence denotes 0.01 M Tris-HCl, pH 7.4/0.01 M NaCl/0.0015 M MgCl,)
TextSentencer_T55 8648-8666 Sentence denotes for 30 min at 4°C.
TextSentencer_T55 8648-8666 Sentence denotes for 30 min at 4°C.
TextSentencer_T56 8667-8746 Sentence denotes The cells were gently Dounce-homogenized and centrifuged at 1000 X g for 5 min.
TextSentencer_T56 8667-8746 Sentence denotes The cells were gently Dounce-homogenized and centrifuged at 1000 X g for 5 min.
TextSentencer_T57 8747-8877 Sentence denotes The nuclear pellet was suspended in the same buffer, Dounce-homogenized and repelleted to increase the purity of this preparation.
TextSentencer_T57 8747-8877 Sentence denotes The nuclear pellet was suspended in the same buffer, Dounce-homogenized and repelleted to increase the purity of this preparation.
TextSentencer_T58 8878-8971 Sentence denotes Following disruption of the nucleus with 0.5% SDS, nucleic acids were extracted as described.
TextSentencer_T58 8878-8971 Sentence denotes Following disruption of the nucleus with 0.5% SDS, nucleic acids were extracted as described.
TextSentencer_T59 8972-9093 Sentence denotes The supernatant, containing membrane and cytosol, was centrifuged at 36000 rpm (100000 x g) in an SW 41 rotor for 90 min.
TextSentencer_T59 8972-9093 Sentence denotes The supernatant, containing membrane and cytosol, was centrifuged at 36000 rpm (100000 x g) in an SW 41 rotor for 90 min.
TextSentencer_T60 9094-9344 Sentence denotes Under these conditions, membranes were not detected in the S-100 fraction by electron microscopy and over 95% of the activities of membrane-associated enzyme markers, e.g. (Na+-K+)-ATPase and 5'-nucleotidase were contained within the P-100 fractions.
TextSentencer_T60 9094-9344 Sentence denotes Under these conditions, membranes were not detected in the S-100 fraction by electron microscopy and over 95% of the activities of membrane-associated enzyme markers, e.g. (Na+-K+)-ATPase and 5'-nucleotidase were contained within the P-100 fractions.
TextSentencer_T61 9345-9465 Sentence denotes The cytosol fraction (S-100) was adjusted to 0.5% SDS and extracted by the phenol/chloroform method (Lai et al., 1981) .
TextSentencer_T61 9345-9465 Sentence denotes The cytosol fraction (S-100) was adjusted to 0.5% SDS and extracted by the phenol/chloroform method (Lai et al., 1981) .
TextSentencer_T62 9466-9605 Sentence denotes The membrane fraction (P-100) was repelleted as described, suspended in 0.5 X RSB containing 0.5% SDS and extracted with phenol/chloroform.
TextSentencer_T62 9466-9605 Sentence denotes The membrane fraction (P-100) was repelleted as described, suspended in 0.5 X RSB containing 0.5% SDS and extracted with phenol/chloroform.
TextSentencer_T63 9606-9794 Sentence denotes Equivalent amounts (25 or 50 pgg/well) of RNA were separated by electrophoresis on 12 or 20% polyacrylamide gels in TBE buffer (0.1 M Tris-borate, pH 8.3/0.002 M EDTA) containing 6 M urea.
TextSentencer_T63 9606-9794 Sentence denotes Equivalent amounts (25 or 50 pgg/well) of RNA were separated by electrophoresis on 12 or 20% polyacrylamide gels in TBE buffer (0.1 M Tris-borate, pH 8.3/0.002 M EDTA) containing 6 M urea.
TextSentencer_T64 9795-9905 Sentence denotes The RNA was electrophoresed at 1000 V until the bromophenol blue marker had migrated 15-20 cm from the origin.
TextSentencer_T64 9795-9905 Sentence denotes The RNA was electrophoresed at 1000 V until the bromophenol blue marker had migrated 15-20 cm from the origin.
TextSentencer_T65 9906-9943 Sentence denotes Under these conditions, intracellular
TextSentencer_T65 9906-9943 Sentence denotes Under these conditions, intracellular
TextSentencer_T66 9944-10044 Sentence denotes RNAs smaller than 250 nucleotides in length enter the gel matrix while the viral mRNAs are excluded.
TextSentencer_T66 9944-10044 Sentence denotes RNAs smaller than 250 nucleotides in length enter the gel matrix while the viral mRNAs are excluded.
TextSentencer_T67 10045-10247 Sentence denotes Urea was removed from the gel matrix by several washings of cold TAE buffer (0.04 M Tris-base/O.02 M Na acetate/O.01 M EDTA, pH 7.4) and the RNA electroblotted to Zeta probe paper (Bio-Rad) in cold TAE.
TextSentencer_T67 10045-10247 Sentence denotes Urea was removed from the gel matrix by several washings of cold TAE buffer (0.04 M Tris-base/O.02 M Na acetate/O.01 M EDTA, pH 7.4) and the RNA electroblotted to Zeta probe paper (Bio-Rad) in cold TAE.
TextSentencer_T68 10248-10325 Sentence denotes Electroblotting was performed in a circulating chamber at 340 mA for lo-15 h.
TextSentencer_T68 10248-10325 Sentence denotes Electroblotting was performed in a circulating chamber at 340 mA for lo-15 h.
TextSentencer_T69 10326-10480 Sentence denotes This protocol efficiently transfers RNA molecules smaller than 250 nucleotides while larger nucleic acids are retained in the gel (Bittner et al., 1980) .
TextSentencer_T69 10326-10480 Sentence denotes This protocol efficiently transfers RNA molecules smaller than 250 nucleotides while larger nucleic acids are retained in the gel (Bittner et al., 1980) .
TextSentencer_T70 10481-10660 Sentence denotes Following transfer, the paper was gently washed in TAE and baked at 80°C for 2 h prior to hybridization with 32P-labeled probes according to published procedures (Thomas, 1980 1 .
TextSentencer_T70 10481-10660 Sentence denotes Following transfer, the paper was gently washed in TAE and baked at 80°C for 2 h prior to hybridization with 32P-labeled probes according to published procedures (Thomas, 1980 1 .
TextSentencer_T71 10661-10755 Sentence denotes Comparison of the junction sequences encoded on genomic RNA. mRNAs 6 and 7 and the leader RNA.
TextSentencer_T71 10661-10755 Sentence denotes Comparison of the junction sequences encoded on genomic RNA. mRNAs 6 and 7 and the leader RNA.
TextSentencer_T72 10756-10933 Sentence denotes The sequences shown are the leader-body junction regions of mRNAs 6 and 7 and the 3'-end sequences of leader RNA (Armstrong et al., 1984; Spaan et al., 1983; Lai et al., 1984) .
TextSentencer_T72 10756-10933 Sentence denotes The sequences shown are the leader-body junction regions of mRNAs 6 and 7 and the 3'-end sequences of leader RNA (Armstrong et al., 1984; Spaan et al., 1983; Lai et al., 1984) .
TextSentencer_T73 10934-11051 Sentence denotes The genomic sequences are from the region which corresponds to the initiation sites for mRNA 7 (Spaan et al., 1983) .
TextSentencer_T73 10934-11051 Sentence denotes The genomic sequences are from the region which corresponds to the initiation sites for mRNA 7 (Spaan et al., 1983) .
TextSentencer_T74 11052-11152 Sentence denotes The sequences underlined represent the common sequences and the possible leader-body junction sites.
TextSentencer_T74 11052-11152 Sentence denotes The sequences underlined represent the common sequences and the possible leader-body junction sites.
TextSentencer_T75 11153-11297 Sentence denotes It has previously been determined that the length of the leader RNA is approximately 72 nucleotides (Armstrong et al., 1984; Lai et al., 1984) .
TextSentencer_T75 11153-11297 Sentence denotes It has previously been determined that the length of the leader RNA is approximately 72 nucleotides (Armstrong et al., 1984; Lai et al., 1984) .
TextSentencer_T76 11298-11630 Sentence denotes This estimate was obtained by primer extension of a synthetic oligodeoxyribonucleotide (5'-AGTT-TAGA'M'AGATT-3') (15-mer), which is complementary to a portion of the splice junction sites of MHV mRNAs 6 and 7 (Lai et al., 1984; Spaan et al., 1983) and also by sequencing of cDNA clones (Armstrong et al., 1984; Spaan et al., 1983) .
TextSentencer_T76 11298-11630 Sentence denotes This estimate was obtained by primer extension of a synthetic oligodeoxyribonucleotide (5'-AGTT-TAGA'M'AGATT-3') (15-mer), which is complementary to a portion of the splice junction sites of MHV mRNAs 6 and 7 (Lai et al., 1984; Spaan et al., 1983) and also by sequencing of cDNA clones (Armstrong et al., 1984; Spaan et al., 1983) .
TextSentencer_T77 11631-11751 Sentence denotes Because of the presence of 15 homologous nucleotides between the 3'-end of the leader RNA and the start site of mRNA No.
TextSentencer_T77 11631-11751 Sentence denotes Because of the presence of 15 homologous nucleotides between the 3'-end of the leader RNA and the start site of mRNA No.
TextSentencer_T78 11752-11964 Sentence denotes 7 on the MHV genome (Fig. f) , it has been suggested that the splice junction lies within a stretch of 15 nucleotides (AAUCUAAUCUAAACU) (nucleotides 58-72 from the 5'-end) (Lai et al., 1984; Spaan et al., 1983) .
TextSentencer_T78 11752-11964 Sentence denotes 7 on the MHV genome (Fig. f) , it has been suggested that the splice junction lies within a stretch of 15 nucleotides (AAUCUAAUCUAAACU) (nucleotides 58-72 from the 5'-end) (Lai et al., 1984; Spaan et al., 1983) .
TextSentencer_T79 11965-12068 Sentence denotes However, the exact length of the leader-coding sequences at the 5'-end of the genomic RNA is not clear.
TextSentencer_T79 11965-12068 Sentence denotes However, the exact length of the leader-coding sequences at the 5'-end of the genomic RNA is not clear.
TextSentencer_T80 12069-12319 Sentence denotes To determine the size of the leader-coding sequences on the genome, we purified the 72-nucleotide leader-specific cDNA made from mRNAs 6 and 7, using the previously described synthetic 15-mer as a primer for reverse transcription (Lai et al., 1984) .
TextSentencer_T80 12069-12319 Sentence denotes To determine the size of the leader-coding sequences on the genome, we purified the 72-nucleotide leader-specific cDNA made from mRNAs 6 and 7, using the previously described synthetic 15-mer as a primer for reverse transcription (Lai et al., 1984) .
TextSentencer_T81 12320-12410 Sentence denotes This leader cDNA was hybridized to purified 60s genomic RNA and digested with S, nuclease.
TextSentencer_T81 12320-12410 Sentence denotes This leader cDNA was hybridized to purified 60s genomic RNA and digested with S, nuclease.
TextSentencer_T82 12411-12484 Sentence denotes No alteration in the size of the 72nucleotide cDNA was observed (Fig 2) .
TextSentencer_T82 12411-12484 Sentence denotes No alteration in the size of the 72nucleotide cDNA was observed (Fig 2) .
TextSentencer_T83 12485-12708 Sentence denotes The minor DNAs seen in Lane A were probably due to random degradation, or the presence of contaminating truncated leader cDNAs, since longer exposure revealed similar patterns in the undigested leader cDNA control (Lane B).
TextSentencer_T83 12485-12708 Sentence denotes The minor DNAs seen in Lane A were probably due to random degradation, or the presence of contaminating truncated leader cDNAs, since longer exposure revealed similar patterns in the undigested leader cDNA control (Lane B).
TextSentencer_T84 12709-12773 Sentence denotes However, minor heterogeneity of genomic RNA cannot be ruled out.
TextSentencer_T84 12709-12773 Sentence denotes However, minor heterogeneity of genomic RNA cannot be ruled out.
TextSentencer_T85 12774-12881 Sentence denotes The leader cDNA was completely digested by S, when it had not been hybridized to the virion RNA (Fig. 2C) .
TextSentencer_T85 12774-12881 Sentence denotes The leader cDNA was completely digested by S, when it had not been hybridized to the virion RNA (Fig. 2C) .
TextSentencer_T86 12882-12978 Sentence denotes These data indicate that a minimum of 72 bases encode the leader RNA at the S-end of the genome.
TextSentencer_T86 12882-12978 Sentence denotes These data indicate that a minimum of 72 bases encode the leader RNA at the S-end of the genome.
TextSentencer_T87 12979-13036 Sentence denotes However, the exact length of the leader RNA is not known.
TextSentencer_T87 12979-13036 Sentence denotes However, the exact length of the leader RNA is not known.
TextSentencer_T88 13037-13178 Sentence denotes We had previously proposed a model of coronavirus mRNA transcription which involved the synthesis of a free leader RNA (Baric et al., 1983) .
TextSentencer_T88 13037-13178 Sentence denotes We had previously proposed a model of coronavirus mRNA transcription which involved the synthesis of a free leader RNA (Baric et al., 1983) .
TextSentencer_T89 13179-13379 Sentence denotes Since the experiments described above indicate that a ~nimum of 72 nucleotides at the 5'-end of the genome encode the leader RNA, the hypothetical free leader RNA synthesized sequences on genomic RNA.
TextSentencer_T89 13179-13379 Sentence denotes Since the experiments described above indicate that a ~nimum of 72 nucleotides at the 5'-end of the genome encode the leader RNA, the hypothetical free leader RNA synthesized sequences on genomic RNA.
TextSentencer_T90 13380-13577 Sentence denotes Internally 32P-labeled leader-specific cDNA (72 nucleotides long) was synthesized from mRNAs 6 and 7 and purified prior to hybridization with genomic RNA under the conditions described in the text.
TextSentencer_T90 13380-13577 Sentence denotes Internally 32P-labeled leader-specific cDNA (72 nucleotides long) was synthesized from mRNAs 6 and 7 and purified prior to hybridization with genomic RNA under the conditions described in the text.
TextSentencer_T91 13578-13698 Sentence denotes After hybridization, the hybrid was digested by S, nuclease and analyzed on 12% polyacrylamide gels containing 6 M urea.
TextSentencer_T91 13578-13698 Sentence denotes After hybridization, the hybrid was digested by S, nuclease and analyzed on 12% polyacrylamide gels containing 6 M urea.
TextSentencer_T92 13699-13739 Sentence denotes Lane A: S-digested leader/genome hybrid.
TextSentencer_T92 13699-13739 Sentence denotes Lane A: S-digested leader/genome hybrid.
TextSentencer_T93 13740-13747 Sentence denotes Lane B:
TextSentencer_T93 13740-13747 Sentence denotes Lane B:
TextSentencer_T94 13748-13767 Sentence denotes Undigested control.
TextSentencer_T94 13748-13767 Sentence denotes Undigested control.
TextSentencer_T95 13768-13775 Sentence denotes Lane C:
TextSentencer_T95 13768-13775 Sentence denotes Lane C:
TextSentencer_T96 13776-13871 Sentence denotes Non-hybridized leader cDNA with S, nuclease. during infection should also be of similar length.
TextSentencer_T96 13776-13871 Sentence denotes Non-hybridized leader cDNA with S, nuclease. during infection should also be of similar length.
TextSentencer_T97 13872-14048 Sentence denotes Initial attempts to identify this hypothetical RNA species, labeled in vivo, were unsuccessful, probably due to high backgrounds of tRNA and other small RNAs in infected cells.
TextSentencer_T97 13872-14048 Sentence denotes Initial attempts to identify this hypothetical RNA species, labeled in vivo, were unsuccessful, probably due to high backgrounds of tRNA and other small RNAs in infected cells.
TextSentencer_T98 14049-14207 Sentence denotes Furthermore, the free leader RNA was probably present in very small quantities since it would be expected to be constantly sequestered to prime transcription.
TextSentencer_T98 14049-14207 Sentence denotes Furthermore, the free leader RNA was probably present in very small quantities since it would be expected to be constantly sequestered to prime transcription.
TextSentencer_T99 14208-14369 Sentence denotes To increase the sensitivity of detection, cDNA specific for the leader sequences was used as a probe to detect the presence of free leader RNA in infected cells.
TextSentencer_T99 14208-14369 Sentence denotes To increase the sensitivity of detection, cDNA specific for the leader sequences was used as a probe to detect the presence of free leader RNA in infected cells.
TextSentencer_T100 14370-14522 Sentence denotes Intracellular RNA was separated on 20% or 12% polyacrylamide gels containing 6 M urea and electroblotted to Zeta-probe paper for Northern blot analysis.
TextSentencer_T100 14370-14522 Sentence denotes Intracellular RNA was separated on 20% or 12% polyacrylamide gels containing 6 M urea and electroblotted to Zeta-probe paper for Northern blot analysis.
TextSentencer_T101 14523-14646 Sentence denotes Under these conditions, nucleic acids smaller than 250 nucleotides in length enter the gel and are transferred efficiently.
TextSentencer_T101 14523-14646 Sentence denotes Under these conditions, nucleic acids smaller than 250 nucleotides in length enter the gel and are transferred efficiently.
TextSentencer_T102 14647-14817 Sentence denotes As shown in Fig. 3A , B, a small RNA species of approximately 75 nucleotides in length was detected with the leader-specific probe in infected, but not uninfected, cells.
TextSentencer_T102 14647-14817 Sentence denotes As shown in Fig. 3A , B, a small RNA species of approximately 75 nucleotides in length was detected with the leader-specific probe in infected, but not uninfected, cells.
TextSentencer_T103 14818-14933 Sentence denotes The size of this leader RNA agrees with that predicted from the S, mapping studies, although it is slightly larger.
TextSentencer_T103 14818-14933 Sentence denotes The size of this leader RNA agrees with that predicted from the S, mapping studies, although it is slightly larger.
TextSentencer_T104 14934-15104 Sentence denotes This result is consistent with the model that the leader RNA is synthesized as a separate entity which is discontinuous from the synthesis of the body sequences of mRNAs.
TextSentencer_T104 14934-15104 Sentence denotes This result is consistent with the model that the leader RNA is synthesized as a separate entity which is discontinuous from the synthesis of the body sequences of mRNAs.
TextSentencer_T105 15105-15264 Sentence denotes In addition to the 75-nucleotide leader RNA species, MHV-infected cells frequently contain multiple leader-related RNA species of different size (Fig. 3A, B) .
TextSentencer_T105 15105-15264 Sentence denotes In addition to the 75-nucleotide leader RNA species, MHV-infected cells frequently contain multiple leader-related RNA species of different size (Fig. 3A, B) .
TextSentencer_T106 15265-15374 Sentence denotes Usually a group of 2-4 species of leader RNAs in the range of 70-82 nucleotides were detected (see also Figs.
TextSentencer_T106 15265-15374 Sentence denotes Usually a group of 2-4 species of leader RNAs in the range of 70-82 nucleotides were detected (see also Figs.
TextSentencer_T107 15375-15385 Sentence denotes 4 and 6) .
TextSentencer_T107 15375-15385 Sentence denotes 4 and 6) .
TextSentencer_T108 15386-15549 Sentence denotes The larger leader-containing RNAs, ranging from approximately 130 to 250 nucleotides in length, were more heterogeneous and varied from preparation to preparation.
TextSentencer_T108 15386-15549 Sentence denotes The larger leader-containing RNAs, ranging from approximately 130 to 250 nucleotides in length, were more heterogeneous and varied from preparation to preparation.
TextSentencer_T109 15550-15810 Sentence denotes Since similar RNA species were not detected by a cDNA probe (clone 143) which represents the first 800 nucleotides from the 3'-end of the genome (unpublished observation) (Fig. 3C) , these data suggest that they were not derived from degradation of viral RNAs.
TextSentencer_T109 15550-15810 Sentence denotes Since similar RNA species were not detected by a cDNA probe (clone 143) which represents the first 800 nucleotides from the 3'-end of the genome (unpublished observation) (Fig. 3C) , these data suggest that they were not derived from degradation of viral RNAs.
TextSentencer_T110 15811-16057 Sentence denotes The presence of these RNAs, which are larger than the 70-82-nucleotide leader RNA, lends further support to a model of mRNA synthesis in which the leader RNA is utilized as a primer, rather than added post-transcriptionally (Baric et al., 1983) .
TextSentencer_T110 15811-16057 Sentence denotes The presence of these RNAs, which are larger than the 70-82-nucleotide leader RNA, lends further support to a model of mRNA synthesis in which the leader RNA is utilized as a primer, rather than added post-transcriptionally (Baric et al., 1983) .
TextSentencer_T111 16058-16199 Sentence denotes The data described above suggests that leader RNA is synthesized as a separate entity which is discontinuous from the synthesis of the mRNAs.
TextSentencer_T111 16058-16199 Sentence denotes The data described above suggests that leader RNA is synthesized as a separate entity which is discontinuous from the synthesis of the mRNAs.
TextSentencer_T112 16200-16399 Sentence denotes It is not clear, however, whether the leader RNA is released from the (-)-strand RNA template into a cytoplasmic pool, or remains tightly complexed with the RNA polymerase and/or (-)-strand template.
TextSentencer_T112 16200-16399 Sentence denotes It is not clear, however, whether the leader RNA is released from the (-)-strand RNA template into a cytoplasmic pool, or remains tightly complexed with the RNA polymerase and/or (-)-strand template.
TextSentencer_T113 16400-16479 Sentence denotes To distinguish between these two possibilities, two approaches were undertaken.
TextSentencer_T113 16400-16479 Sentence denotes To distinguish between these two possibilities, two approaches were undertaken.
TextSentencer_T114 16480-16560 Sentence denotes The first was to determine the subcellular localization of the leader-containing
TextSentencer_T114 16480-16560 Sentence denotes The first was to determine the subcellular localization of the leader-containing
TextSentencer_T115 16561-16566 Sentence denotes RNAs.
TextSentencer_T115 16561-16566 Sentence denotes RNAs.
TextSentencer_T116 16567-16728 Sentence denotes It has previously been shown that RNA synthesis takes place in membrane-associated complexes during MHV infection (Brayton et al., 1982 (Brayton et al., , 1984 .
TextSentencer_T116 16567-16728 Sentence denotes It has previously been shown that RNA synthesis takes place in membrane-associated complexes during MHV infection (Brayton et al., 1982 (Brayton et al., , 1984 .
TextSentencer_T117 16729-16890 Sentence denotes Therefore, distribution of the leader RNA in different subcellular compartments should elucidate the mechanism of transcription involving the leader RNA species.
TextSentencer_T117 16729-16890 Sentence denotes Therefore, distribution of the leader RNA in different subcellular compartments should elucidate the mechanism of transcription involving the leader RNA species.
TextSentencer_T118 16891-17025 Sentence denotes MHV-infected cells were disrupted and separated into nuclear, cytosol (S-100) and membrane (P-100) fractions (Stohlman et al., 1983) .
TextSentencer_T118 16891-17025 Sentence denotes MHV-infected cells were disrupted and separated into nuclear, cytosol (S-100) and membrane (P-100) fractions (Stohlman et al., 1983) .
TextSentencer_T119 17026-17141 Sentence denotes RNA from each fraction was separated on polyacrylamide gels and probed for leader-specific RNA sequences (Fig. 4) .
TextSentencer_T119 17026-17141 Sentence denotes RNA from each fraction was separated on polyacrylamide gels and probed for leader-specific RNA sequences (Fig. 4) .
TextSentencer_T120 17142-17281 Sentence denotes All of the leader-containing RNAs were present in both the membrane and cytosol fractions while none were detected in the nuclear fraction.
TextSentencer_T120 17142-17281 Sentence denotes All of the leader-containing RNAs were present in both the membrane and cytosol fractions while none were detected in the nuclear fraction.
TextSentencer_T121 17282-17546 Sentence denotes It is noteworthy that the leader RNAs in the range of 70-82 nucleotides were detected in roughly equimolar amounts in the cytosol and membrane fractions while the larger leader-related RNAs (greater than 130 nucleotides) were detected predominantly in the cytosol.
TextSentencer_T121 17282-17546 Sentence denotes It is noteworthy that the leader RNAs in the range of 70-82 nucleotides were detected in roughly equimolar amounts in the cytosol and membrane fractions while the larger leader-related RNAs (greater than 130 nucleotides) were detected predominantly in the cytosol.
TextSentencer_T122 17547-17755 Sentence denotes This result is consistent with the interpretation that part of the 70-82-nucleotide leader RNA species is coupled to the membrane-associated transcriptional complex and involved in the synthesis of each mRNA.
TextSentencer_T122 17547-17755 Sentence denotes This result is consistent with the interpretation that part of the 70-82-nucleotide leader RNA species is coupled to the membrane-associated transcriptional complex and involved in the synthesis of each mRNA.
TextSentencer_T123 17756-18048 Sentence denotes Moreover, although we can not completely rule out the possibility that our fractionation procedures might have disrupted the replication complexes, the detection of a significant portion of these RNA species in cytosol suggests that the leader RNA might also be present in a cytoplasmic pool.
TextSentencer_T123 17756-18048 Sentence denotes Moreover, although we can not completely rule out the possibility that our fractionation procedures might have disrupted the replication complexes, the detection of a significant portion of these RNA species in cytosol suggests that the leader RNA might also be present in a cytoplasmic pool.
TextSentencer_T124 18049-18219 Sentence denotes We have studied the effects of temperature at which the virus is grown, with the purpose of further uncoupling the synthesis of leader-containing RNAs from that of mRNAs.
TextSentencer_T124 18049-18219 Sentence denotes We have studied the effects of temperature at which the virus is grown, with the purpose of further uncoupling the synthesis of leader-containing RNAs from that of mRNAs.
TextSentencer_T125 18220-18350 Sentence denotes Infected cells maintained at 40°C (Lane 4 in Fig. 4 ) appear to contain less leader RNA species than those grown at 37°C (Lane 5).
TextSentencer_T125 18220-18350 Sentence denotes Infected cells maintained at 40°C (Lane 4 in Fig. 4 ) appear to contain less leader RNA species than those grown at 37°C (Lane 5).
TextSentencer_T126 18351-18470 Sentence denotes This difference roughly corresponded to the general levels of RNA synthesis at these two temperatures (data not shown).
TextSentencer_T126 18351-18470 Sentence denotes This difference roughly corresponded to the general levels of RNA synthesis at these two temperatures (data not shown).
TextSentencer_T127 18471-18577 Sentence denotes Therefore, the synthesis of leader RNA and that of mRNAs could not be uncoupled at these two temperatures.
TextSentencer_T127 18471-18577 Sentence denotes Therefore, the synthesis of leader RNA and that of mRNAs could not be uncoupled at these two temperatures.
TextSentencer_T128 18578-18836 Sentence denotes To determine whether the leader-containing RNAs are associated with the RNA template, intracellular RNA from infected cells was extracted and separated by sucrose gradient sedimentation to determine the size of RNA fractions in which leader RNA was detected.
TextSentencer_T128 18578-18836 Sentence denotes To determine whether the leader-containing RNAs are associated with the RNA template, intracellular RNA from infected cells was extracted and separated by sucrose gradient sedimentation to determine the size of RNA fractions in which leader RNA was detected.
TextSentencer_T129 18837-18921 Sentence denotes The majority of the leader-containing RNAs were found in 4S RNA fractions (Fig. 5) .
TextSentencer_T129 18837-18921 Sentence denotes The majority of the leader-containing RNAs were found in 4S RNA fractions (Fig. 5) .
TextSentencer_T130 18922-19181 Sentence denotes Since the larger-size RNA fractions do not contain distinct leader-related RNA species (Fig. 5) , these results suggest that at least some leader RNA species are dissociated from the RNA replicative intermediate, which is 12-328 in size (Baric et al., 1983) .
TextSentencer_T130 18922-19181 Sentence denotes Since the larger-size RNA fractions do not contain distinct leader-related RNA species (Fig. 5) , these results suggest that at least some leader RNA species are dissociated from the RNA replicative intermediate, which is 12-328 in size (Baric et al., 1983) .
TextSentencer_T131 19182-19349 Sentence denotes However, this conclusion has to be qualified since The cells were disrupted and separated into nuclear, cytoplasmic (S-100) or membrane (P-100) fractions as described.
TextSentencer_T131 19182-19349 Sentence denotes However, this conclusion has to be qualified since The cells were disrupted and separated into nuclear, cytoplasmic (S-100) or membrane (P-100) fractions as described.
TextSentencer_T132 19350-19461 Sentence denotes Intracellular RNA was isolated from each fraction and separated on 12% polyacrylamide gels containing 6 M urea.
TextSentencer_T132 19350-19461 Sentence denotes Intracellular RNA was isolated from each fraction and separated on 12% polyacrylamide gels containing 6 M urea.
TextSentencer_T133 19462-19566 Sentence denotes Following transfer to Zeta-probe paper, the blots were hybridized with 32P-labeled leader-specific cDNA.
TextSentencer_T133 19462-19566 Sentence denotes Following transfer to Zeta-probe paper, the blots were hybridized with 32P-labeled leader-specific cDNA.
TextSentencer_T134 19567-19574 Sentence denotes Lane 1:
TextSentencer_T134 19567-19574 Sentence denotes Lane 1:
TextSentencer_T135 19575-19601 Sentence denotes Purified nuclear fraction.
TextSentencer_T135 19575-19601 Sentence denotes Purified nuclear fraction.
TextSentencer_T136 19602-19609 Sentence denotes Lane 2:
TextSentencer_T136 19602-19609 Sentence denotes Lane 2:
TextSentencer_T137 19610-19644 Sentence denotes Purified membrane fraction, P-100.
TextSentencer_T137 19610-19644 Sentence denotes Purified membrane fraction, P-100.
TextSentencer_T138 19645-19652 Sentence denotes Lane 3:
TextSentencer_T138 19645-19652 Sentence denotes Lane 3:
TextSentencer_T139 19653-19678 Sentence denotes Purified cytosoi (S-100).
TextSentencer_T139 19653-19678 Sentence denotes Purified cytosoi (S-100).
TextSentencer_T140 19679-19714 Sentence denotes Lane 4: A59-infected ceils at 40°C.
TextSentencer_T140 19679-19714 Sentence denotes Lane 4: A59-infected ceils at 40°C.
TextSentencer_T141 19715-19750 Sentence denotes Lane 5: A59-infected cells at 37°C.
TextSentencer_T141 19715-19750 Sentence denotes Lane 5: A59-infected cells at 37°C.
TextSentencer_T142 19751-19758 Sentence denotes Lane 6:
TextSentencer_T142 19751-19758 Sentence denotes Lane 6:
TextSentencer_T143 19759-19803 Sentence denotes In vitro synthesized leader cDNA as markers.
TextSentencer_T143 19759-19803 Sentence denotes In vitro synthesized leader cDNA as markers.
TextSentencer_T144 19804-19851 Sentence denotes The arrow denotes the 7Znucleotide leader cDNA.
TextSentencer_T144 19804-19851 Sentence denotes The arrow denotes the 7Znucleotide leader cDNA.
TextSentencer_T145 19852-20002 Sentence denotes we can not rule out the possibility that the leader RNA might have been dissociated from the replicative intermediate during the extraction procedure.
TextSentencer_T145 19852-20002 Sentence denotes we can not rule out the possibility that the leader RNA might have been dissociated from the replicative intermediate during the extraction procedure.
TextSentencer_T146 20003-20201 Sentence denotes These data are consistent with the subcellular fractionation studies which suggest that the leader-related RNAs are synthesized as a free entity and may function in viral transcription on membranes.
TextSentencer_T146 20003-20201 Sentence denotes These data are consistent with the subcellular fractionation studies which suggest that the leader-related RNAs are synthesized as a free entity and may function in viral transcription on membranes.
TextSentencer_T147 20202-20396 Sentence denotes The detection of free leader-containing RNAs in MHV-infected cells provides support for a mechanism of coronavirus transcription which utilizes a small leader RNA as a primer for mRNA synthesis.
TextSentencer_T147 20202-20396 Sentence denotes The detection of free leader-containing RNAs in MHV-infected cells provides support for a mechanism of coronavirus transcription which utilizes a small leader RNA as a primer for mRNA synthesis.
TextSentencer_T148 20397-20555 Sentence denotes However, definitive proof of a distinct leader-specific RNA is complicated by the presence of multiple small leader-containing RNAs in infected cell extracts.
TextSentencer_T148 20397-20555 Sentence denotes However, definitive proof of a distinct leader-specific RNA is complicated by the presence of multiple small leader-containing RNAs in infected cell extracts.
TextSentencer_T149 20556-20754 Sentence denotes To provide additional evidence for leader-primed trans~~ption, we reasoned that the synthesis of leader RNA and mRNAs should be discontinuous and, therefore, might require different viral functions.
TextSentencer_T149 20556-20754 Sentence denotes To provide additional evidence for leader-primed trans~~ption, we reasoned that the synthesis of leader RNA and mRNAs should be discontinuous and, therefore, might require different viral functions.
TextSentencer_T150 20755-20853 Sentence denotes Thus, it might and sedimented at 25 000 rpm on lo-25% sucrose gradients for 16 h in an SW41 rotor.
TextSentencer_T150 20755-20853 Sentence denotes Thus, it might and sedimented at 25 000 rpm on lo-25% sucrose gradients for 16 h in an SW41 rotor.
TextSentencer_T151 20854-20889 Sentence denotes The gradients were pooled into 28s.
TextSentencer_T151 20854-20889 Sentence denotes The gradients were pooled into 28s.
TextSentencer_T152 20890-20983 Sentence denotes 18s and 4S fractions (a) and electrophoresed on polyacrylamide gels as described in the text.
TextSentencer_T152 20890-20983 Sentence denotes 18s and 4S fractions (a) and electrophoresed on polyacrylamide gels as described in the text.
TextSentencer_T153 20984-21080 Sentence denotes Following transfer, the blotting paper was probed with the 32P-labeled leader-specific cDNA (b).
TextSentencer_T153 20984-21080 Sentence denotes Following transfer, the blotting paper was probed with the 32P-labeled leader-specific cDNA (b).
TextSentencer_T154 21081-21134 Sentence denotes Lanes A-C represent fractions I to III, respectively.
TextSentencer_T154 21081-21134 Sentence denotes Lanes A-C represent fractions I to III, respectively.
TextSentencer_T155 21135-21203 Sentence denotes The arrow denotes the in vitro synthesized 72nucleotide leader cDNA.
TextSentencer_T155 21135-21203 Sentence denotes The arrow denotes the in vitro synthesized 72nucleotide leader cDNA.
TextSentencer_T156 21204-21311 Sentence denotes be possible to isolate a temperature-sensitive (ts) mutant which synthesizes only leader RNA but not mRNAs.
TextSentencer_T156 21204-21311 Sentence denotes be possible to isolate a temperature-sensitive (ts) mutant which synthesizes only leader RNA but not mRNAs.
TextSentencer_T157 21312-21432 Sentence denotes We therefore examined several ts mutants of MHV for their ability to synthesize leader RNA at nonpermissive temperature.
TextSentencer_T157 21312-21432 Sentence denotes We therefore examined several ts mutants of MHV for their ability to synthesize leader RNA at nonpermissive temperature.
TextSentencer_T158 21433-21532 Sentence denotes The majority of these ts mutants are RNA-( -), and fail to synthesize mRNAs later in the infection.
TextSentencer_T158 21433-21532 Sentence denotes The majority of these ts mutants are RNA-( -), and fail to synthesize mRNAs later in the infection.
TextSentencer_T159 21533-21654 Sentence denotes These mutants also grow significantly more slowly than the wild-type virus at nonpermissive temperature (data not shown).
TextSentencer_T159 21533-21654 Sentence denotes These mutants also grow significantly more slowly than the wild-type virus at nonpermissive temperature (data not shown).
TextSentencer_T160 21655-21799 Sentence denotes Cells were infected with different ts mutants and incubated at permissive temperature for the first 6 h to permit synthesis of (-)-stranded RNA.
TextSentencer_T160 21655-21799 Sentence denotes Cells were infected with different ts mutants and incubated at permissive temperature for the first 6 h to permit synthesis of (-)-stranded RNA.
TextSentencer_T161 21800-21956 Sentence denotes The cultures were then shifted to nonpermissive temperature prior to the onset ( + )-strand synthesis and analyzed for the presence of leader RNA 4 h later.
TextSentencer_T161 21800-21956 Sentence denotes The cultures were then shifted to nonpermissive temperature prior to the onset ( + )-strand synthesis and analyzed for the presence of leader RNA 4 h later.
TextSentencer_T162 21957-22074 Sentence denotes Most of these ts mutants failed to synthesize any detectable RNA when probed with the leader-specific cDNA (Fig. 6) .
TextSentencer_T162 21957-22074 Sentence denotes Most of these ts mutants failed to synthesize any detectable RNA when probed with the leader-specific cDNA (Fig. 6) .
TextSentencer_T163 22075-22213 Sentence denotes This is consistent with the observation that these mutants fail to synthesize mRNA at nonpermissive temperature (unpublished observation).
TextSentencer_T163 22075-22213 Sentence denotes This is consistent with the observation that these mutants fail to synthesize mRNA at nonpermissive temperature (unpublished observation).
TextSentencer_T164 22214-22331 Sentence denotes However, one mutant, LAlO, synthesizes a series of small leader-related RNA species at the nonpermissive temperature.
TextSentencer_T164 22214-22331 Sentence denotes However, one mutant, LAlO, synthesizes a series of small leader-related RNA species at the nonpermissive temperature.
TextSentencer_T165 22332-22475 Sentence denotes The largest RNA had a similar size to the 72-nucleotide leader cDNA marker, although several smaller leader-containing RNAs were also detected.
TextSentencer_T165 22332-22475 Sentence denotes The largest RNA had a similar size to the 72-nucleotide leader cDNA marker, although several smaller leader-containing RNAs were also detected.
TextSentencer_T166 22476-22526 Sentence denotes Since we did not detect larger leader-re- Fig. 6 .
TextSentencer_T166 22476-22526 Sentence denotes Since we did not detect larger leader-re- Fig. 6 .
TextSentencer_T167 22527-22611 Sentence denotes Analysis of leader RNA synthesis with A59 ts mutants at the restrictive temperature.
TextSentencer_T167 22527-22611 Sentence denotes Analysis of leader RNA synthesis with A59 ts mutants at the restrictive temperature.
TextSentencer_T168 22612-22683 Sentence denotes Cultures of L2 cells were infected with a variety of ts mutants of A59.
TextSentencer_T168 22612-22683 Sentence denotes Cultures of L2 cells were infected with a variety of ts mutants of A59.
TextSentencer_T169 22684-22841 Sentence denotes Following 1 h for virus adsorption, the medium was replaced and the cultures incubated at 32°C for 6 h prior to shift-up to 39°C (nonpermissive temperature).
TextSentencer_T169 22684-22841 Sentence denotes Following 1 h for virus adsorption, the medium was replaced and the cultures incubated at 32°C for 6 h prior to shift-up to 39°C (nonpermissive temperature).
TextSentencer_T170 22842-22945 Sentence denotes Intracellular RNA was extracted at 10 h post-infection, and electrophoresed on 20% polyacrylamide gels.
TextSentencer_T170 22842-22945 Sentence denotes Intracellular RNA was extracted at 10 h post-infection, and electrophoresed on 20% polyacrylamide gels.
TextSentencer_T171 22946-23037 Sentence denotes After electrotransfer to Zeta probe paper, the blots were probed with leader-specific cDNA.
TextSentencer_T171 22946-23037 Sentence denotes After electrotransfer to Zeta probe paper, the blots were probed with leader-specific cDNA.
TextSentencer_T172 23038-23058 Sentence denotes Lanes l-7: LAl, LA2.
TextSentencer_T172 23038-23058 Sentence denotes Lanes l-7: LAl, LA2.
TextSentencer_T173 23059-23084 Sentence denotes LA5, LA6, LA8, LAlO, XA9.
TextSentencer_T173 23059-23084 Sentence denotes LA5, LA6, LA8, LAlO, XA9.
TextSentencer_T174 23085-23133 Sentence denotes The arrow denotes the 72-nucleotide leader cDNA.
TextSentencer_T174 23085-23133 Sentence denotes The arrow denotes the 72-nucleotide leader cDNA.
TextSentencer_T175 23134-23276 Sentence denotes lated RNAs, these data indicate that the replication of this mutant is blocked at a step after the synthesis of this 72-nucleotide leader RNA.
TextSentencer_T175 23134-23276 Sentence denotes lated RNAs, these data indicate that the replication of this mutant is blocked at a step after the synthesis of this 72-nucleotide leader RNA.
TextSentencer_T176 23277-23442 Sentence denotes They also suggest that this RNA species is not a premature termination product of mRNA synthesis, and that its synthesis is discontinuous from the synthesis of mRNA.
TextSentencer_T176 23277-23442 Sentence denotes They also suggest that this RNA species is not a premature termination product of mRNA synthesis, and that its synthesis is discontinuous from the synthesis of mRNA.
TextSentencer_T177 23443-23582 Sentence denotes The phenotype of LA10 is RNA-( -), as determined by rates of RNA synthesis at permissive and nonpermissive temperatures later in infection.
TextSentencer_T177 23443-23582 Sentence denotes The phenotype of LA10 is RNA-( -), as determined by rates of RNA synthesis at permissive and nonpermissive temperatures later in infection.
TextSentencer_T178 23583-23714 Sentence denotes Temperature-shift experiments also demonstrated that this mutant is defective in early steps of viral replication (data not shown).
TextSentencer_T178 23583-23714 Sentence denotes Temperature-shift experiments also demonstrated that this mutant is defective in early steps of viral replication (data not shown).
TextSentencer_T179 23715-23922 Sentence denotes We have also examined an RNA-( +) mutant, XA9, which synthesizes the larger leader-containing RNA species, identical to those observed for wild-type virus, at the nonpermissive temperature (Fig. 6, lane 7) .
TextSentencer_T179 23715-23922 Sentence denotes We have also examined an RNA-( +) mutant, XA9, which synthesizes the larger leader-containing RNA species, identical to those observed for wild-type virus, at the nonpermissive temperature (Fig. 6, lane 7) .
TextSentencer_T180 23923-23995 Sentence denotes These results are consistent with the RNA-( +) phenotype of this mutant.
TextSentencer_T180 23923-23995 Sentence denotes These results are consistent with the RNA-( +) phenotype of this mutant.
TextSentencer_T181 23996-24108 Sentence denotes These results strongly suggest that the synthesis of the leader RNA and mRNAs requires separate viral functions.
TextSentencer_T181 23996-24108 Sentence denotes These results strongly suggest that the synthesis of the leader RNA and mRNAs requires separate viral functions.
TextSentencer_T182 24109-24342 Sentence denotes The presence of a mutant which accumulates leader-specific RNA at nonpermissive temperature lends further support to the model that the leader RNA is synthesized independently and then utilized as a primer for the synthesis of mRNAs.
TextSentencer_T182 24109-24342 Sentence denotes The presence of a mutant which accumulates leader-specific RNA at nonpermissive temperature lends further support to the model that the leader RNA is synthesized independently and then utilized as a primer for the synthesis of mRNAs.
TextSentencer_T183 24343-24577 Sentence denotes Previous studies on MHV mRNA structure indicate that each viral niRNA is constructed by the joining of two noncontiguous RNA segments encoded on the genome (Baric et al., 1983; Lai et al., 1983 Lai et al., , 1984 Spaan et al., 1983) .
TextSentencer_T183 24343-24577 Sentence denotes Previous studies on MHV mRNA structure indicate that each viral niRNA is constructed by the joining of two noncontiguous RNA segments encoded on the genome (Baric et al., 1983; Lai et al., 1983 Lai et al., , 1984 Spaan et al., 1983) .
TextSentencer_T184 24578-24880 Sentence denotes Since MHV mRNA synthesis occurs in the cytoplasm and does not involve synthesis and subsequent cleavage of precursor RNAs, viral transcription must occur by a novel mechanism that differs from conventional splicing modes (Baric et al., 1983; Lai et al., 1983; Jacobs et al., 1981; Spaan et al., 1983) .
TextSentencer_T184 24578-24880 Sentence denotes Since MHV mRNA synthesis occurs in the cytoplasm and does not involve synthesis and subsequent cleavage of precursor RNAs, viral transcription must occur by a novel mechanism that differs from conventional splicing modes (Baric et al., 1983; Lai et al., 1983; Jacobs et al., 1981; Spaan et al., 1983) .
TextSentencer_T185 24881-25099 Sentence denotes Analysis of MHV replicative intermediate and replicative form RNAs supports this contention and suggests that a free leader RNA is transcribed independently, and then used to prime mRNA synthesis (Baric et al., 1983) .
TextSentencer_T185 24881-25099 Sentence denotes Analysis of MHV replicative intermediate and replicative form RNAs supports this contention and suggests that a free leader RNA is transcribed independently, and then used to prime mRNA synthesis (Baric et al., 1983) .
TextSentencer_T186 25100-25212 Sentence denotes In this publication, we have demonstrated the presence of multiple leader-containing RNAs in MHV-infected cells.
TextSentencer_T186 25100-25212 Sentence denotes In this publication, we have demonstrated the presence of multiple leader-containing RNAs in MHV-infected cells.
TextSentencer_T187 25213-25322 Sentence denotes Furthermore, we have identified a ts mutant which accumulates small leader RNAs at nonpermissive temperature.
TextSentencer_T187 25213-25322 Sentence denotes Furthermore, we have identified a ts mutant which accumulates small leader RNAs at nonpermissive temperature.
TextSentencer_T188 25323-25583 Sentence denotes These findings provide strong evidence that MHV leader RNA synthesis is separate from the transcription of the mRNAs and provide additional support for the leader-primed mechanism of coronavirus transcription which we proposed previously (Baric et al., 1983) .
TextSentencer_T188 25323-25583 Sentence denotes These findings provide strong evidence that MHV leader RNA synthesis is separate from the transcription of the mRNAs and provide additional support for the leader-primed mechanism of coronavirus transcription which we proposed previously (Baric et al., 1983) .
TextSentencer_T189 25584-25821 Sentence denotes Sequence analysis of mRNAs 7 and 6 suggests that the junction sites between the leader RNA and the body sequences of each mRNA lie between nucleotides 58 and 72 from the 5'-end of the genome (Armstrong et al., 1984; Spaan et al., 1983) .
TextSentencer_T189 25584-25821 Sentence denotes Sequence analysis of mRNAs 7 and 6 suggests that the junction sites between the leader RNA and the body sequences of each mRNA lie between nucleotides 58 and 72 from the 5'-end of the genome (Armstrong et al., 1984; Spaan et al., 1983) .
TextSentencer_T190 25822-25979 Sentence denotes From S, mapping studies, the length of the leader-coding sequences at the 5'-end of the genome is at least 72 nucleotides, but the exact length is not known.
TextSentencer_T190 25822-25979 Sentence denotes From S, mapping studies, the length of the leader-coding sequences at the 5'-end of the genome is at least 72 nucleotides, but the exact length is not known.
TextSentencer_T191 25980-26093 Sentence denotes The major free leader RNA species identified in the MHV-infected cells is approximately 75 nucleotides in length.
TextSentencer_T191 25980-26093 Sentence denotes The major free leader RNA species identified in the MHV-infected cells is approximately 75 nucleotides in length.
TextSentencer_T192 26094-26251 Sentence denotes However, these data were complicated by the presence of several additional discrete leader-specific RNAs which range between 70 and 82 nucleotides in length.
TextSentencer_T192 26094-26251 Sentence denotes However, these data were complicated by the presence of several additional discrete leader-specific RNAs which range between 70 and 82 nucleotides in length.
TextSentencer_T193 26252-26323 Sentence denotes Several explanations could account for the multiple leaderrelated RNAs.
TextSentencer_T193 26252-26323 Sentence denotes Several explanations could account for the multiple leaderrelated RNAs.
TextSentencer_T194 26324-26510 Sentence denotes Analysis of the primary sequence of the leader RNA suggests that several double-stranded RNA loops, similar to tRNA structure, may be present in the leader RNA (unpublished observation).
TextSentencer_T194 26324-26510 Sentence denotes Analysis of the primary sequence of the leader RNA suggests that several double-stranded RNA loops, similar to tRNA structure, may be present in the leader RNA (unpublished observation).
TextSentencer_T195 26511-26647 Sentence denotes These loops could represent strong stops for RNA synthesis, causing the accumulation of discrete premature termination products in vivo.
TextSentencer_T195 26511-26647 Sentence denotes These loops could represent strong stops for RNA synthesis, causing the accumulation of discrete premature termination products in vivo.
TextSentencer_T196 26648-26753 Sentence denotes Alternatively, the different leader RNAs might be primers for transcription of the different viral mRNAs.
TextSentencer_T196 26648-26753 Sentence denotes Alternatively, the different leader RNAs might be primers for transcription of the different viral mRNAs.
TextSentencer_T197 26754-27030 Sentence denotes It has previously been suggested that differences in the degree of complementarity between the 3'-end of leader RNA and different junction sequences on the (-)-strand could regulate the rate of synthesis of each mRNA species (Fig. 1) (Baric et al., 1983; Spaan et al., 1983) .
TextSentencer_T197 26754-27030 Sentence denotes It has previously been suggested that differences in the degree of complementarity between the 3'-end of leader RNA and different junction sequences on the (-)-strand could regulate the rate of synthesis of each mRNA species (Fig. 1) (Baric et al., 1983; Spaan et al., 1983) .
TextSentencer_T198 27031-27178 Sentence denotes The requirement for different leader RNAs might also provide an additional mechanism for the regulation of transcription of different mRNA species.
TextSentencer_T198 27031-27178 Sentence denotes The requirement for different leader RNAs might also provide an additional mechanism for the regulation of transcription of different mRNA species.
TextSentencer_T199 27179-27427 Sentence denotes The leader-primed transcription model is further supported by our recent finding that the leader sequences could be exchanged at very high frequency between two different MHV strains during mixed infection (Makino et al., unpublished observation) .
TextSentencer_T199 27179-27427 Sentence denotes The leader-primed transcription model is further supported by our recent finding that the leader sequences could be exchanged at very high frequency between two different MHV strains during mixed infection (Makino et al., unpublished observation) .
TextSentencer_T200 27428-27528 Sentence denotes These data provide strong evidence for a mechanism of MHV transcription involving a free leader RNA.
TextSentencer_T200 27428-27528 Sentence denotes These data provide strong evidence for a mechanism of MHV transcription involving a free leader RNA.
TextSentencer_T201 27529-27632 Sentence denotes However, definitive proof that these leader RNAs act as primers awaits an in vitro transcription assay.
TextSentencer_T201 27529-27632 Sentence denotes However, definitive proof that these leader RNAs act as primers awaits an in vitro transcription assay.
TextSentencer_T202 27633-27800 Sentence denotes The larger leader-related RNA species which are longer than 130 nucleotides might represent intermediate RNA products caused by premature termination of transcription.
TextSentencer_T202 27633-27800 Sentence denotes The larger leader-related RNA species which are longer than 130 nucleotides might represent intermediate RNA products caused by premature termination of transcription.
TextSentencer_T203 27801-28012 Sentence denotes The fact that these RNA species contain the leader sequences and are significantly larger than the 75nucleotide leader RNA provides further evidence that the leader RNA is utilized as a primer for transcription.
TextSentencer_T203 27801-28012 Sentence denotes The fact that these RNA species contain the leader sequences and are significantly larger than the 75nucleotide leader RNA provides further evidence that the leader RNA is utilized as a primer for transcription.
TextSentencer_T204 28013-28060 Sentence denotes It is conceivable that these intermediate-sized
TextSentencer_T204 28013-28060 Sentence denotes It is conceivable that these intermediate-sized
TextSentencer_T205 28061-28130 Sentence denotes leader-related RNAs also participate in the process of transcription,
TextSentencer_T205 28061-28130 Sentence denotes leader-related RNAs also participate in the process of transcription,
TextSentencer_T206 28131-28212 Sentence denotes i.e. they can bind back to the RNA template and act as primers for transcription.
TextSentencer_T206 28131-28212 Sentence denotes i.e. they can bind back to the RNA template and act as primers for transcription.
TextSentencer_T207 28213-28379 Sentence denotes This possibility has recently been supported by the isolation of recombinant RNA molecules during mixed infection of different MHV strains (unpublished observations).
TextSentencer_T207 28213-28379 Sentence denotes This possibility has recently been supported by the isolation of recombinant RNA molecules during mixed infection of different MHV strains (unpublished observations).
TextSentencer_T208 28380-28552 Sentence denotes The uncoupling of leader RNA synthesis from mRNA synthesis with ts-LA10 provides strong evidence that transcription of MHV mRNAs requires more than one viral gene function.
TextSentencer_T208 28380-28552 Sentence denotes The uncoupling of leader RNA synthesis from mRNA synthesis with ts-LA10 provides strong evidence that transcription of MHV mRNAs requires more than one viral gene function.
TextSentencer_T209 28553-28676 Sentence denotes It is possible that two different polymerases are responsible for the synthesis of the leader RNAs and mRNAs, respectively.
TextSentencer_T209 28553-28676 Sentence denotes It is possible that two different polymerases are responsible for the synthesis of the leader RNAs and mRNAs, respectively.
TextSentencer_T210 28677-28864 Sentence denotes Alternatively, a specific viral protein might be required to either bind the leader RNA to the RNA template or modify RNA polymerase to allow the transcription of body sequences of mRNAs.
TextSentencer_T210 28677-28864 Sentence denotes Alternatively, a specific viral protein might be required to either bind the leader RNA to the RNA template or modify RNA polymerase to allow the transcription of body sequences of mRNAs.
TextSentencer_T211 28865-28996 Sentence denotes Subunit regulation of a core polymerase has been proposed for the synthesis of alphavirus subgenomic mRNAs (Sawicki et al., 1978) .
TextSentencer_T211 28865-28996 Sentence denotes Subunit regulation of a core polymerase has been proposed for the synthesis of alphavirus subgenomic mRNAs (Sawicki et al., 1978) .
TextSentencer_T212 28997-29086 Sentence denotes The 'leader-primed' mechanism for MHV RNA synthesis is a feature unique to coronaviruses.
TextSentencer_T212 28997-29086 Sentence denotes The 'leader-primed' mechanism for MHV RNA synthesis is a feature unique to coronaviruses.
TextSentencer_T213 29087-29234 Sentence denotes Several other viruses, such as VSV, a rhabdovirus, and La Crosse virus, a bunyavirus, also synthesize free leader RNAs (Colonno and Banerjee, 1976:
TextSentencer_T213 29087-29234 Sentence denotes Several other viruses, such as VSV, a rhabdovirus, and La Crosse virus, a bunyavirus, also synthesize free leader RNAs (Colonno and Banerjee, 1976:
TextSentencer_T214 29235-29260 Sentence denotes Patterson et al., 1983) .
TextSentencer_T214 29235-29260 Sentence denotes Patterson et al., 1983) .
TextSentencer_T215 29261-29327 Sentence denotes However, these leader RNAs are not joined covalently to the mRNAs.
TextSentencer_T215 29261-29327 Sentence denotes However, these leader RNAs are not joined covalently to the mRNAs.
TextSentencer_T216 29328-29552 Sentence denotes While the function of the bunyavirus leader RNAs has not been determined, the 47-nucleotide VSV leader is transported to the nucleus and serves as an inhibitor of host transcription (Grinell and Wagner, 1984; Kurilla et al..
TextSentencer_T216 29328-29552 Sentence denotes While the function of the bunyavirus leader RNAs has not been determined, the 47-nucleotide VSV leader is transported to the nucleus and serves as an inhibitor of host transcription (Grinell and Wagner, 1984; Kurilla et al..
TextSentencer_T217 29553-29582 Sentence denotes 1982; McGowan et al., 1982) .
TextSentencer_T217 29553-29582 Sentence denotes 1982; McGowan et al., 1982) .
TextSentencer_T218 29583-29652 Sentence denotes Similar functions have not been detected with coronavirus leader RNA.
TextSentencer_T218 29583-29652 Sentence denotes Similar functions have not been detected with coronavirus leader RNA.
TextSentencer_T219 29653-29749 Sentence denotes On the other hand, the transcription of coronavirus partially resembles that of influenza virus.
TextSentencer_T219 29653-29749 Sentence denotes On the other hand, the transcription of coronavirus partially resembles that of influenza virus.
TextSentencer_T220 29750-29929 Sentence denotes Although influenza virus does not synthesize a free leader RNA, it requires a host-derived 5'-capped RNA as a primer for transcription (Bouloy et al., 1978; Plotch et al., 1981) .
TextSentencer_T220 29750-29929 Sentence denotes Although influenza virus does not synthesize a free leader RNA, it requires a host-derived 5'-capped RNA as a primer for transcription (Bouloy et al., 1978; Plotch et al., 1981) .
TextSentencer_T221 29930-30046 Sentence denotes The leader-primed mechanism for RNA transcription employed by coronaviruses might have even more general occurrence.
TextSentencer_T221 29930-30046 Sentence denotes The leader-primed mechanism for RNA transcription employed by coronaviruses might have even more general occurrence.
TextSentencer_T222 30047-30259 Sentence denotes It has been shown that the mRNAs of the variable surface glycoprotein (VSG) in African trypanosomes contain an identical 37-nucleotide stretch at their 5'-ends (Boothroyd and Cross, 1982; Longacre et al., 1983) .
TextSentencer_T222 30047-30259 Sentence denotes It has been shown that the mRNAs of the variable surface glycoprotein (VSG) in African trypanosomes contain an identical 37-nucleotide stretch at their 5'-ends (Boothroyd and Cross, 1982; Longacre et al., 1983) .
TextSentencer_T223 30260-30427 Sentence denotes More recent studies have shown that a free leader RNA is synthesized which might serve as the primer for the VSG mRNA (Campbell et al., 1984; Milhausen et al., 1984) .
TextSentencer_T223 30260-30427 Sentence denotes More recent studies have shown that a free leader RNA is synthesized which might serve as the primer for the VSG mRNA (Campbell et al., 1984; Milhausen et al., 1984) .
TextSentencer_T224 30428-30571 Sentence denotes Thus, this leader-primed RNA transcription might represent a common alternative mechanism for generating spliced mRNAs in eukaryotic organisms.
TextSentencer_T224 30428-30571 Sentence denotes Thus, this leader-primed RNA transcription might represent a common alternative mechanism for generating spliced mRNAs in eukaryotic organisms.