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pred":"punct","subj":"T7534","obj":"T7522"},{"id":"R4892","pred":"nmod","subj":"T7536","obj":"T7537"},{"id":"R4893","pred":"nsubj","subj":"T7537","obj":"T7541"},{"id":"R4894","pred":"punct","subj":"T7538","obj":"T7536"},{"id":"R4895","pred":"punct","subj":"T7539","obj":"T7536"},{"id":"R4896","pred":"punct","subj":"T7540","obj":"T7536"},{"id":"R4897","pred":"det","subj":"T7542","obj":"T7543"},{"id":"R4898","pred":"dobj","subj":"T7543","obj":"T7541"},{"id":"R4899","pred":"amod","subj":"T7544","obj":"T7543"},{"id":"R4900","pred":"prep","subj":"T7545","obj":"T7541"},{"id":"R4901","pred":"det","subj":"T7546","obj":"T7547"},{"id":"R4902","pred":"pobj","subj":"T7547","obj":"T7545"},{"id":"R4903","pred":"acl","subj":"T7548","obj":"T7547"},{"id":"R4904","pred":"punct","subj":"T7549","obj":"T7541"}],"text":"RNA Analysis\nRT-PCR amplification from exon 7 to 11 from total heart RNA amplified the expected 600-base pair (bp) fragment in MCAD+/+ and MCAD+/− mice, and a 1.5-kb fragment in MCAD−/− mice (data not shown). Sequence analysis of the 1.5-kb fragment revealed that the amplified fragment consisted of exon 7 to exon 10 with 280 bp of pGEM plasmid sequence followed by exons 8–11. Some of the plasmid sequences, along with the pPGKNEOpA sequence, were deleted from this spliced mRNA.\nNorthern blot analysis revealed Acadm was normally expressed in all tissues analyzed from MCAD+/+ mice with the most robust expression occurring in brown fat, kidney, heart, skeletal muscle, and liver with minimal expression in the brain, white fat, and testes (Figure 2). Interestingly, although RT-PCR amplified an incorrectly spliced Acadm transcript, no Acadm transcripts were detected by northern blot analysis from MCAD−/− mice. These results strongly suggest that the mutant RNA is unstable and degraded rapidly or, alternatively, undergoes nonsense mediated RNA decay.\nFigure 2 Northern Blot Analysis from MCAD−/− (n = 2) and MCAD+/+ (n = 2) Mice\nAcadm message was detected from the heart, liver, brown fat, brain, kidney, and muscle (and white fat and testes, data not shown) of only MCAD+/+ mice. Most robust expression occurred in brown fat, kidney, heart, and skeletal muscle. MCAD−/− mice had no detectable message in all tissues examined."}

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Sequence analysis of the 1.5-kb fragment revealed that the amplified fragment consisted of exon 7 to exon 10 with 280 bp of pGEM plasmid sequence followed by exons 8–11. Some of the plasmid sequences, along with the pPGKNEOpA sequence, were deleted from this spliced mRNA.\nNorthern blot analysis revealed Acadm was normally expressed in all tissues analyzed from MCAD+/+ mice with the most robust expression occurring in brown fat, kidney, heart, skeletal muscle, and liver with minimal expression in the brain, white fat, and testes (Figure 2). Interestingly, although RT-PCR amplified an incorrectly spliced Acadm transcript, no Acadm transcripts were detected by northern blot analysis from MCAD−/− mice. These results strongly suggest that the mutant RNA is unstable and degraded rapidly or, alternatively, undergoes nonsense mediated RNA decay.\nFigure 2 Northern Blot Analysis from MCAD−/− (n = 2) and MCAD+/+ (n = 2) Mice\nAcadm message was detected from the heart, liver, brown fat, brain, kidney, and muscle (and white fat and testes, data not shown) of only MCAD+/+ mice. Most robust expression occurred in brown fat, kidney, heart, and skeletal muscle. MCAD−/− mice had no detectable message in all tissues examined."}

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Analysis\nRT-PCR amplification from exon 7 to 11 from total heart RNA amplified the expected 600-base pair (bp) fragment in MCAD+/+ and MCAD+/− mice, and a 1.5-kb fragment in MCAD−/− mice (data not shown). Sequence analysis of the 1.5-kb fragment revealed that the amplified fragment consisted of exon 7 to exon 10 with 280 bp of pGEM plasmid sequence followed by exons 8–11. Some of the plasmid sequences, along with the pPGKNEOpA sequence, were deleted from this spliced mRNA.\nNorthern blot analysis revealed Acadm was normally expressed in all tissues analyzed from MCAD+/+ mice with the most robust expression occurring in brown fat, kidney, heart, skeletal muscle, and liver with minimal expression in the brain, white fat, and testes (Figure 2). Interestingly, although RT-PCR amplified an incorrectly spliced Acadm transcript, no Acadm transcripts were detected by northern blot analysis from MCAD−/− mice. These results strongly suggest that the mutant RNA is unstable and degraded rapidly or, alternatively, undergoes nonsense mediated RNA decay.\nFigure 2 Northern Blot Analysis from MCAD−/− (n = 2) and MCAD+/+ (n = 2) Mice\nAcadm message was detected from the heart, liver, brown fat, brain, kidney, and muscle (and white fat and testes, data not shown) of only MCAD+/+ mice. Most robust expression occurred in brown fat, kidney, heart, and skeletal muscle. MCAD−/− mice had no detectable message in all tissues examined."}