Id |
Subject |
Object |
Predicate |
Lexical cue |
T4611 |
0-9 |
NN |
denotes |
Targeting |
T4612 |
10-19 |
NN |
denotes |
construct |
T4613 |
20-23 |
IN |
denotes |
for |
T4614 |
24-25 |
DT |
denotes |
a |
T4615 |
41-46 |
NN |
denotes |
locus |
T4616 |
26-29 |
NN |
denotes |
p53 |
T4617 |
30-34 |
NN |
denotes |
RMCE |
T4618 |
35-40 |
JJ |
denotes |
ready |
T4619 |
34-35 |
HYPH |
denotes |
- |
T4620 |
46-160 |
sentence |
denotes |
Details for plasmid construction are available upon request owing to space limitations mandating a brief outline. |
T4621 |
47-54 |
NNS |
denotes |
Details |
T4622 |
80-83 |
VBP |
denotes |
are |
T4623 |
55-58 |
IN |
denotes |
for |
T4624 |
59-66 |
NN |
denotes |
plasmid |
T4625 |
67-79 |
NN |
denotes |
construction |
T4626 |
84-93 |
JJ |
denotes |
available |
T4627 |
94-98 |
IN |
denotes |
upon |
T4628 |
99-106 |
NN |
denotes |
request |
T4629 |
107-112 |
VBG |
denotes |
owing |
T4630 |
113-115 |
IN |
denotes |
to |
T4631 |
116-121 |
NN |
denotes |
space |
T4632 |
122-133 |
NNS |
denotes |
limitations |
T4633 |
134-143 |
VBG |
denotes |
mandating |
T4634 |
144-145 |
DT |
denotes |
a |
T4635 |
152-159 |
NN |
denotes |
outline |
T4636 |
146-151 |
JJ |
denotes |
brief |
T4637 |
159-160 |
. |
denotes |
. |
T4638 |
160-306 |
sentence |
denotes |
We started from a plasmid L3-1L containing heterologous loxP sites (L3 is the mutant loxP257 recently described (14), 1L is an inverted WT loxP). |
T4639 |
161-163 |
PRP |
denotes |
We |
T4640 |
164-171 |
VBD |
denotes |
started |
T4641 |
172-176 |
IN |
denotes |
from |
T4642 |
177-178 |
DT |
denotes |
a |
T4643 |
190-192 |
NN |
denotes |
1L |
T4644 |
179-186 |
NN |
denotes |
plasmid |
T4645 |
187-189 |
NN |
denotes |
L3 |
T4646 |
189-190 |
HYPH |
denotes |
- |
T4647 |
193-203 |
VBG |
denotes |
containing |
T4648 |
204-216 |
JJ |
denotes |
heterologous |
T4649 |
222-227 |
NNS |
denotes |
sites |
T4650 |
217-221 |
NN |
denotes |
loxP |
T4651 |
228-229 |
-LRB- |
denotes |
( |
T4652 |
282-284 |
VBZ |
denotes |
is |
T4653 |
229-231 |
NN |
denotes |
L3 |
T4654 |
232-234 |
VBZ |
denotes |
is |
T4655 |
235-238 |
DT |
denotes |
the |
T4656 |
246-253 |
NN |
denotes |
loxP257 |
T4657 |
239-245 |
JJ |
denotes |
mutant |
T4658 |
254-262 |
RB |
denotes |
recently |
T4659 |
263-272 |
VBN |
denotes |
described |
T4660 |
273-274 |
-LRB- |
denotes |
( |
T4661 |
274-276 |
CD |
denotes |
14 |
T4662 |
276-277 |
-RRB- |
denotes |
) |
T4663 |
277-279 |
, |
denotes |
, |
T4664 |
279-281 |
NN |
denotes |
1L |
T4665 |
285-287 |
DT |
denotes |
an |
T4666 |
300-304 |
NN |
denotes |
loxP |
T4667 |
288-296 |
VBN |
denotes |
inverted |
T4668 |
297-299 |
NN |
denotes |
WT |
T4669 |
304-305 |
-RRB- |
denotes |
) |
T4670 |
305-306 |
. |
denotes |
. |
T4671 |
306-446 |
sentence |
denotes |
The WT loxP and loxP257 differ in their spacer sequences: the spacer sequence is 5′-ATGTATGC-3′ for WT loxP and 5′-AAGTCTCC-3′ for loxP257. |
T4672 |
307-310 |
DT |
denotes |
The |
T4673 |
314-318 |
NN |
denotes |
loxP |
T4674 |
311-313 |
NN |
denotes |
WT |
T4675 |
331-337 |
VBP |
denotes |
differ |
T4676 |
319-322 |
CC |
denotes |
and |
T4677 |
323-330 |
NN |
denotes |
loxP257 |
T4678 |
385-387 |
VBZ |
denotes |
is |
T4679 |
338-340 |
IN |
denotes |
in |
T4680 |
341-346 |
PRP$ |
denotes |
their |
T4681 |
354-363 |
NNS |
denotes |
sequences |
T4682 |
347-353 |
NN |
denotes |
spacer |
T4683 |
363-365 |
: |
denotes |
: |
T4684 |
365-368 |
DT |
denotes |
the |
T4685 |
376-384 |
NN |
denotes |
sequence |
T4686 |
369-375 |
NN |
denotes |
spacer |
T4687 |
388-389 |
CD |
denotes |
5 |
T4688 |
389-390 |
SYM |
denotes |
′ |
T4689 |
390-391 |
HYPH |
denotes |
- |
T4690 |
391-399 |
JJ |
denotes |
ATGTATGC |
T4691 |
399-400 |
HYPH |
denotes |
- |
T4692 |
400-401 |
CD |
denotes |
3 |
T4693 |
401-402 |
SYM |
denotes |
′ |
T4694 |
403-406 |
IN |
denotes |
for |
T4695 |
407-409 |
NN |
denotes |
WT |
T4696 |
410-414 |
NN |
denotes |
loxP |
T4697 |
415-418 |
CC |
denotes |
and |
T4698 |
419-420 |
CD |
denotes |
5 |
T4699 |
422-430 |
NN |
denotes |
AAGTCTCC |
T4700 |
420-421 |
SYM |
denotes |
′ |
T4701 |
421-422 |
HYPH |
denotes |
- |
T4702 |
430-431 |
HYPH |
denotes |
- |
T4703 |
431-432 |
CD |
denotes |
3 |
T4704 |
432-433 |
SYM |
denotes |
′ |
T4705 |
434-437 |
IN |
denotes |
for |
T4706 |
438-445 |
NN |
denotes |
loxP257 |
T4707 |
445-446 |
. |
denotes |
. |
T4708 |
446-750 |
sentence |
denotes |
The three mutations in the loxP257 spacer sequence prevent it to recombine with WT loxP, ensuring efficient RMCE in several cell lines: accurate RMCE with these loxP sites occurred with an average frequency of 81% at two loci in CHO cells and an average frequency of 69% at four loci in Hela cells (14). |
T4709 |
447-450 |
DT |
denotes |
The |
T4710 |
457-466 |
NNS |
denotes |
mutations |
T4711 |
451-456 |
CD |
denotes |
three |
T4712 |
498-505 |
VBP |
denotes |
prevent |
T4713 |
467-469 |
IN |
denotes |
in |
T4714 |
470-473 |
DT |
denotes |
the |
T4715 |
489-497 |
NN |
denotes |
sequence |
T4716 |
474-481 |
NN |
denotes |
loxP257 |
T4717 |
482-488 |
NN |
denotes |
spacer |
T4718 |
619-627 |
VBD |
denotes |
occurred |
T4719 |
506-508 |
PRP |
denotes |
it |
T4720 |
512-521 |
VB |
denotes |
recombine |
T4721 |
509-511 |
TO |
denotes |
to |
T4722 |
522-526 |
IN |
denotes |
with |
T4723 |
527-529 |
NN |
denotes |
WT |
T4724 |
530-534 |
NN |
denotes |
loxP |
T4725 |
534-536 |
, |
denotes |
, |
T4726 |
536-544 |
VBG |
denotes |
ensuring |
T4727 |
545-554 |
JJ |
denotes |
efficient |
T4728 |
555-559 |
NN |
denotes |
RMCE |
T4729 |
560-562 |
IN |
denotes |
in |
T4730 |
563-570 |
JJ |
denotes |
several |
T4731 |
576-581 |
NNS |
denotes |
lines |
T4732 |
571-575 |
NN |
denotes |
cell |
T4733 |
581-583 |
: |
denotes |
: |
T4734 |
583-591 |
JJ |
denotes |
accurate |
T4735 |
592-596 |
NN |
denotes |
RMCE |
T4736 |
597-601 |
IN |
denotes |
with |
T4737 |
602-607 |
DT |
denotes |
these |
T4738 |
613-618 |
NNS |
denotes |
sites |
T4739 |
608-612 |
NN |
denotes |
loxP |
T4740 |
628-632 |
IN |
denotes |
with |
T4741 |
633-635 |
DT |
denotes |
an |
T4742 |
644-653 |
NN |
denotes |
frequency |
T4743 |
636-643 |
JJ |
denotes |
average |
T4744 |
654-656 |
IN |
denotes |
of |
T4745 |
657-659 |
CD |
denotes |
81 |
T4746 |
659-660 |
NN |
denotes |
% |
T4747 |
661-663 |
IN |
denotes |
at |
T4748 |
664-667 |
CD |
denotes |
two |
T4749 |
668-672 |
NNS |
denotes |
loci |
T4750 |
673-675 |
IN |
denotes |
in |
T4751 |
676-679 |
NN |
denotes |
CHO |
T4752 |
680-685 |
NNS |
denotes |
cells |
T4753 |
686-689 |
CC |
denotes |
and |
T4754 |
690-692 |
DT |
denotes |
an |
T4755 |
701-710 |
NN |
denotes |
frequency |
T4756 |
693-700 |
JJ |
denotes |
average |
T4757 |
711-713 |
IN |
denotes |
of |
T4758 |
714-716 |
CD |
denotes |
69 |
T4759 |
716-717 |
NN |
denotes |
% |
T4760 |
718-720 |
IN |
denotes |
at |
T4761 |
721-725 |
CD |
denotes |
four |
T4762 |
726-730 |
NNS |
denotes |
loci |
T4763 |
731-733 |
IN |
denotes |
in |
T4764 |
734-738 |
NN |
denotes |
Hela |
T4765 |
739-744 |
NNS |
denotes |
cells |
T4766 |
745-746 |
-LRB- |
denotes |
( |
T4767 |
746-748 |
CD |
denotes |
14 |
T4768 |
748-749 |
-RRB- |
denotes |
) |
T4769 |
749-750 |
. |
denotes |
. |
T4770 |
750-874 |
sentence |
denotes |
The L3-1L plasmid was first modified to include a ClaI and a FseI site between the LoxP sites, leading to plasmid L3-CF-1L. |
T4771 |
751-754 |
DT |
denotes |
The |
T4772 |
761-768 |
NN |
denotes |
plasmid |
T4773 |
755-757 |
NN |
denotes |
L3 |
T4774 |
758-760 |
NN |
denotes |
1L |
T4775 |
757-758 |
HYPH |
denotes |
- |
T4776 |
779-787 |
VBN |
denotes |
modified |
T4777 |
769-772 |
VBD |
denotes |
was |
T4778 |
773-778 |
RB |
denotes |
first |
T4779 |
788-790 |
TO |
denotes |
to |
T4780 |
791-798 |
VB |
denotes |
include |
T4781 |
799-800 |
DT |
denotes |
a |
T4782 |
801-805 |
NN |
denotes |
ClaI |
T4783 |
806-809 |
CC |
denotes |
and |
T4784 |
810-811 |
DT |
denotes |
a |
T4785 |
817-821 |
NN |
denotes |
site |
T4786 |
812-816 |
NN |
denotes |
FseI |
T4787 |
822-829 |
IN |
denotes |
between |
T4788 |
830-833 |
DT |
denotes |
the |
T4789 |
839-844 |
NNS |
denotes |
sites |
T4790 |
834-838 |
NN |
denotes |
LoxP |
T4791 |
844-846 |
, |
denotes |
, |
T4792 |
846-853 |
VBG |
denotes |
leading |
T4793 |
854-856 |
IN |
denotes |
to |
T4794 |
857-864 |
NN |
denotes |
plasmid |
T4795 |
871-873 |
NN |
denotes |
1L |
T4796 |
865-867 |
NN |
denotes |
L3 |
T4797 |
867-868 |
HYPH |
denotes |
- |
T4798 |
868-870 |
NN |
denotes |
CF |
T4799 |
870-871 |
HYPH |
denotes |
- |
T4800 |
873-874 |
. |
denotes |
. |
T4801 |
874-1136 |
sentence |
denotes |
We next modified a puroΔTK plasmid (YTC37, a kind gift from A. Bradley) by using oligonucleotides to destroy a NotI site downstream of the puroΔTK gene and introduce a NotI site upstream, and a FseI site downstream of the gene (leading to plasmid CN-PuroΔTK-F). |
T4802 |
875-877 |
PRP |
denotes |
We |
T4803 |
883-891 |
VBD |
denotes |
modified |
T4804 |
878-882 |
RB |
denotes |
next |
T4805 |
892-893 |
DT |
denotes |
a |
T4806 |
902-909 |
NN |
denotes |
plasmid |
T4807 |
894-901 |
NN |
denotes |
puroΔTK |
T4808 |
910-911 |
-LRB- |
denotes |
( |
T4809 |
911-916 |
NN |
denotes |
YTC37 |
T4810 |
916-918 |
, |
denotes |
, |
T4811 |
918-919 |
DT |
denotes |
a |
T4812 |
925-929 |
NN |
denotes |
gift |
T4813 |
920-924 |
NN |
denotes |
kind |
T4814 |
930-934 |
IN |
denotes |
from |
T4815 |
935-937 |
NNP |
denotes |
A. |
T4816 |
938-945 |
NNP |
denotes |
Bradley |
T4817 |
945-946 |
-RRB- |
denotes |
) |
T4818 |
947-949 |
IN |
denotes |
by |
T4819 |
950-955 |
VBG |
denotes |
using |
T4820 |
956-972 |
NNS |
denotes |
oligonucleotides |
T4821 |
973-975 |
TO |
denotes |
to |
T4822 |
976-983 |
VB |
denotes |
destroy |
T4823 |
984-985 |
DT |
denotes |
a |
T4824 |
991-995 |
NN |
denotes |
site |
T4825 |
986-990 |
NN |
denotes |
NotI |
T4826 |
996-1006 |
RB |
denotes |
downstream |
T4827 |
1007-1009 |
IN |
denotes |
of |
T4828 |
1010-1013 |
DT |
denotes |
the |
T4829 |
1022-1026 |
NN |
denotes |
gene |
T4830 |
1014-1021 |
NN |
denotes |
puroΔTK |
T4831 |
1027-1030 |
CC |
denotes |
and |
T4832 |
1031-1040 |
VB |
denotes |
introduce |
T4833 |
1041-1042 |
DT |
denotes |
a |
T4834 |
1048-1052 |
NN |
denotes |
site |
T4835 |
1043-1047 |
NN |
denotes |
NotI |
T4836 |
1053-1061 |
RB |
denotes |
upstream |
T4837 |
1061-1063 |
, |
denotes |
, |
T4838 |
1063-1066 |
CC |
denotes |
and |
T4839 |
1067-1068 |
DT |
denotes |
a |
T4840 |
1074-1078 |
NN |
denotes |
site |
T4841 |
1069-1073 |
NN |
denotes |
FseI |
T4842 |
1079-1089 |
RB |
denotes |
downstream |
T4843 |
1090-1092 |
IN |
denotes |
of |
T4844 |
1093-1096 |
DT |
denotes |
the |
T4845 |
1097-1101 |
NN |
denotes |
gene |
T4846 |
1102-1103 |
-LRB- |
denotes |
( |
T4847 |
1103-1110 |
VBG |
denotes |
leading |
T4848 |
1111-1113 |
IN |
denotes |
to |
T4849 |
1114-1121 |
NN |
denotes |
plasmid |
T4850 |
1133-1134 |
NN |
denotes |
F |
T4851 |
1122-1124 |
NN |
denotes |
CN |
T4852 |
1124-1125 |
HYPH |
denotes |
- |
T4853 |
1125-1132 |
NN |
denotes |
PuroΔTK |
T4854 |
1132-1133 |
HYPH |
denotes |
- |
T4855 |
1134-1135 |
-RRB- |
denotes |
) |
T4856 |
1135-1136 |
. |
denotes |
. |
T4857 |
1136-1384 |
sentence |
denotes |
Next, a PmlI-MfeI 6.3 kb fragment from Trp53 was subcloned in a modified pBluescript KSII+ (pBS, Stratagene), and the resulting plasmid was digested with SwaI to introduce an EagI site, leading to p53PmlEag, a plasmid containing exons 2–11 of p53. |
T4858 |
1137-1141 |
RB |
denotes |
Next |
T4859 |
1186-1195 |
VBN |
denotes |
subcloned |
T4860 |
1141-1143 |
, |
denotes |
, |
T4861 |
1143-1144 |
DT |
denotes |
a |
T4862 |
1162-1170 |
NN |
denotes |
fragment |
T4863 |
1145-1149 |
NN |
denotes |
PmlI |
T4864 |
1150-1154 |
NN |
denotes |
MfeI |
T4865 |
1149-1150 |
HYPH |
denotes |
- |
T4866 |
1155-1158 |
CD |
denotes |
6.3 |
T4867 |
1159-1161 |
NN |
denotes |
kb |
T4868 |
1171-1175 |
IN |
denotes |
from |
T4869 |
1176-1181 |
NN |
denotes |
Trp53 |
T4870 |
1182-1185 |
VBD |
denotes |
was |
T4871 |
1196-1198 |
IN |
denotes |
in |
T4872 |
1199-1200 |
DT |
denotes |
a |
T4873 |
1222-1227 |
NN |
denotes |
KSII+ |
T4874 |
1201-1209 |
VBN |
denotes |
modified |
T4875 |
1210-1221 |
NN |
denotes |
pBluescript |
T4876 |
1228-1229 |
-LRB- |
denotes |
( |
T4877 |
1229-1232 |
NN |
denotes |
pBS |
T4878 |
1232-1234 |
, |
denotes |
, |
T4879 |
1234-1244 |
NNP |
denotes |
Stratagene |
T4880 |
1244-1245 |
-RRB- |
denotes |
) |
T4881 |
1245-1247 |
, |
denotes |
, |
T4882 |
1247-1250 |
CC |
denotes |
and |
T4883 |
1251-1254 |
DT |
denotes |
the |
T4884 |
1265-1272 |
NN |
denotes |
plasmid |
T4885 |
1255-1264 |
VBG |
denotes |
resulting |
T4886 |
1277-1285 |
VBN |
denotes |
digested |
T4887 |
1273-1276 |
VBD |
denotes |
was |
T4888 |
1286-1290 |
IN |
denotes |
with |
T4889 |
1291-1295 |
NN |
denotes |
SwaI |
T4890 |
1296-1298 |
TO |
denotes |
to |
T4891 |
1299-1308 |
VB |
denotes |
introduce |
T4892 |
1309-1311 |
DT |
denotes |
an |
T4893 |
1317-1321 |
NN |
denotes |
site |
T4894 |
1312-1316 |
NN |
denotes |
EagI |
T4895 |
1321-1323 |
, |
denotes |
, |
T4896 |
1323-1330 |
VBG |
denotes |
leading |
T4897 |
1331-1333 |
IN |
denotes |
to |
T4898 |
1334-1343 |
NN |
denotes |
p53PmlEag |
T4899 |
1343-1345 |
, |
denotes |
, |
T4900 |
1345-1346 |
DT |
denotes |
a |
T4901 |
1347-1354 |
NN |
denotes |
plasmid |
T4902 |
1355-1365 |
VBG |
denotes |
containing |
T4903 |
1366-1371 |
NNS |
denotes |
exons |
T4904 |
1374-1376 |
CD |
denotes |
11 |
T4905 |
1372-1373 |
CD |
denotes |
2 |
T4906 |
1373-1374 |
HYPH |
denotes |
– |
T4907 |
1377-1379 |
IN |
denotes |
of |
T4908 |
1380-1383 |
NN |
denotes |
p53 |
T4909 |
1383-1384 |
. |
denotes |
. |
T4910 |
1384-1629 |
sentence |
denotes |
We then inserted a 5.5 kb ClaI-EagI fragment from p53PmlEag in plasmid CN-PuroΔTK-F digested by ClaI and NotI, and inserted the resulting fragment between the loxP sites of L3-CF-1L by ClaI and FseI digestion, leading to L3-p53PmlEagPuroΔTK-1L. |
T4911 |
1385-1387 |
PRP |
denotes |
We |
T4912 |
1393-1401 |
VBD |
denotes |
inserted |
T4913 |
1388-1392 |
RB |
denotes |
then |
T4914 |
1402-1403 |
DT |
denotes |
a |
T4915 |
1421-1429 |
NN |
denotes |
fragment |
T4916 |
1404-1407 |
CD |
denotes |
5.5 |
T4917 |
1408-1410 |
NN |
denotes |
kb |
T4918 |
1411-1415 |
NN |
denotes |
ClaI |
T4919 |
1416-1420 |
NN |
denotes |
EagI |
T4920 |
1415-1416 |
HYPH |
denotes |
- |
T4921 |
1430-1434 |
IN |
denotes |
from |
T4922 |
1435-1444 |
NN |
denotes |
p53PmlEag |
T4923 |
1445-1447 |
IN |
denotes |
in |
T4924 |
1448-1455 |
NN |
denotes |
plasmid |
T4925 |
1467-1468 |
NN |
denotes |
F |
T4926 |
1456-1458 |
NN |
denotes |
CN |
T4927 |
1458-1459 |
HYPH |
denotes |
- |
T4928 |
1459-1466 |
NN |
denotes |
PuroΔTK |
T4929 |
1466-1467 |
HYPH |
denotes |
- |
T4930 |
1469-1477 |
VBN |
denotes |
digested |
T4931 |
1478-1480 |
IN |
denotes |
by |
T4932 |
1481-1485 |
NN |
denotes |
ClaI |
T4933 |
1486-1489 |
CC |
denotes |
and |
T4934 |
1490-1494 |
NN |
denotes |
NotI |
T4935 |
1494-1496 |
, |
denotes |
, |
T4936 |
1496-1499 |
CC |
denotes |
and |
T4937 |
1500-1508 |
VBD |
denotes |
inserted |
T4938 |
1509-1512 |
DT |
denotes |
the |
T4939 |
1523-1531 |
NN |
denotes |
fragment |
T4940 |
1513-1522 |
VBG |
denotes |
resulting |
T4941 |
1532-1539 |
IN |
denotes |
between |
T4942 |
1540-1543 |
DT |
denotes |
the |
T4943 |
1549-1554 |
NNS |
denotes |
sites |
T4944 |
1544-1548 |
NN |
denotes |
loxP |
T4945 |
1555-1557 |
IN |
denotes |
of |
T4946 |
1558-1560 |
NN |
denotes |
L3 |
T4947 |
1564-1566 |
NN |
denotes |
1L |
T4948 |
1560-1561 |
HYPH |
denotes |
- |
T4949 |
1561-1563 |
NN |
denotes |
CF |
T4950 |
1563-1564 |
HYPH |
denotes |
- |
T4951 |
1567-1569 |
IN |
denotes |
by |
T4952 |
1570-1574 |
NN |
denotes |
ClaI |
T4953 |
1584-1593 |
NN |
denotes |
digestion |
T4954 |
1575-1578 |
CC |
denotes |
and |
T4955 |
1579-1583 |
NN |
denotes |
FseI |
T4956 |
1593-1595 |
, |
denotes |
, |
T4957 |
1595-1602 |
VBG |
denotes |
leading |
T4958 |
1603-1605 |
IN |
denotes |
to |
T4959 |
1606-1608 |
NN |
denotes |
L3 |
T4960 |
1626-1628 |
NN |
denotes |
1L |
T4961 |
1608-1609 |
HYPH |
denotes |
- |
T4962 |
1609-1625 |
NN |
denotes |
p53PmlEagPuroΔTK |
T4963 |
1625-1626 |
HYPH |
denotes |
- |
T4964 |
1628-1629 |
. |
denotes |
. |
T4965 |
1629-2219 |
sentence |
denotes |
We next engineered a plasmid containing the region for 3′-homology downstream of the p53 gene and the DTA gene in two steps: (i) we performed a three-way ligation between a modified pBS digested by HindIII and NotI, a HindIII-EcoRI fragment from Trp53 for 3′homology and an EcoRI-NotI fragment containing the DTA gene, from plasmid pgkdtabpa (kind gift of P. Soriano), leading to plasmid 3′ + DTA and (ii) because the Bsu36I-EcoRI region downstream of p53 contains repetitive sequences (F. Toledo and G. M. Wahl, unpublished data), we later deleted this region, to obtain plasmid 3′ + DTA. |
T4966 |
1630-1632 |
PRP |
denotes |
We |
T4967 |
1638-1648 |
VBD |
denotes |
engineered |
T4968 |
1633-1637 |
RB |
denotes |
next |
T4969 |
1762-1771 |
VBD |
denotes |
performed |
T4970 |
1649-1650 |
DT |
denotes |
a |
T4971 |
1651-1658 |
NN |
denotes |
plasmid |
T4972 |
1659-1669 |
VBG |
denotes |
containing |
T4973 |
1670-1673 |
DT |
denotes |
the |
T4974 |
1674-1680 |
NN |
denotes |
region |
T4975 |
1681-1684 |
IN |
denotes |
for |
T4976 |
1685-1686 |
CD |
denotes |
3 |
T4977 |
1686-1687 |
SYM |
denotes |
′ |
T4978 |
1687-1688 |
SYM |
denotes |
- |
T4979 |
1688-1696 |
JJ |
denotes |
homology |
T4980 |
1697-1707 |
RB |
denotes |
downstream |
T4981 |
1708-1710 |
IN |
denotes |
of |
T4982 |
1711-1714 |
DT |
denotes |
the |
T4983 |
1719-1723 |
NN |
denotes |
gene |
T4984 |
1715-1718 |
NN |
denotes |
p53 |
T4985 |
1724-1727 |
CC |
denotes |
and |
T4986 |
1728-1731 |
DT |
denotes |
the |
T4987 |
1736-1740 |
NN |
denotes |
gene |
T4988 |
1732-1735 |
NN |
denotes |
DTA |
T4989 |
1741-1743 |
IN |
denotes |
in |
T4990 |
1744-1747 |
CD |
denotes |
two |
T4991 |
1748-1753 |
NNS |
denotes |
steps |
T4992 |
1753-1755 |
: |
denotes |
: |
T4993 |
1755-1756 |
-LRB- |
denotes |
( |
T4994 |
1756-1757 |
LS |
denotes |
i |
T4995 |
1757-1758 |
-RRB- |
denotes |
) |
T4996 |
1759-1761 |
PRP |
denotes |
we |
T4997 |
1772-1773 |
DT |
denotes |
a |
T4998 |
1784-1792 |
NN |
denotes |
ligation |
T4999 |
1774-1779 |
CD |
denotes |
three |
T5000 |
1780-1783 |
NN |
denotes |
way |
T5001 |
1779-1780 |
HYPH |
denotes |
- |
T5002 |
1793-1800 |
IN |
denotes |
between |
T5003 |
1801-1802 |
DT |
denotes |
a |
T5004 |
1812-1815 |
NN |
denotes |
pBS |
T5005 |
1803-1811 |
VBN |
denotes |
modified |
T5006 |
1816-1824 |
VBN |
denotes |
digested |
T5007 |
1825-1827 |
IN |
denotes |
by |
T5008 |
1828-1835 |
NN |
denotes |
HindIII |
T5009 |
1836-1839 |
CC |
denotes |
and |
T5010 |
1840-1844 |
NN |
denotes |
NotI |
T5011 |
1844-1846 |
, |
denotes |
, |
T5012 |
1846-1847 |
DT |
denotes |
a |
T5013 |
1862-1870 |
NN |
denotes |
fragment |
T5014 |
1848-1855 |
NN |
denotes |
HindIII |
T5015 |
1856-1861 |
NN |
denotes |
EcoRI |
T5016 |
1855-1856 |
HYPH |
denotes |
- |
T5017 |
1871-1875 |
IN |
denotes |
from |
T5018 |
1876-1881 |
NN |
denotes |
Trp53 |
T5019 |
1882-1885 |
IN |
denotes |
for |
T5020 |
1886-1887 |
CD |
denotes |
3 |
T5021 |
1888-1896 |
NN |
denotes |
homology |
T5022 |
1887-1888 |
SYM |
denotes |
′ |
T5023 |
1897-1900 |
CC |
denotes |
and |
T5024 |
1901-1903 |
DT |
denotes |
an |
T5025 |
1915-1923 |
NN |
denotes |
fragment |
T5026 |
1904-1909 |
NN |
denotes |
EcoRI |
T5027 |
1910-1914 |
NN |
denotes |
NotI |
T5028 |
1909-1910 |
HYPH |
denotes |
- |
T5029 |
1924-1934 |
VBG |
denotes |
containing |
T5030 |
1935-1938 |
DT |
denotes |
the |
T5031 |
1943-1947 |
NN |
denotes |
gene |
T5032 |
1939-1942 |
NN |
denotes |
DTA |
T5033 |
1947-1949 |
, |
denotes |
, |
T5034 |
1949-1953 |
IN |
denotes |
from |
T5035 |
1954-1961 |
NN |
denotes |
plasmid |
T5036 |
1962-1971 |
NN |
denotes |
pgkdtabpa |
T5037 |
1972-1973 |
-LRB- |
denotes |
( |
T5038 |
1973-1977 |
JJ |
denotes |
kind |
T5039 |
1978-1982 |
NN |
denotes |
gift |
T5040 |
1983-1985 |
IN |
denotes |
of |
T5041 |
1986-1988 |
NNP |
denotes |
P. |
T5042 |
1989-1996 |
NNP |
denotes |
Soriano |
T5043 |
1996-1997 |
-RRB- |
denotes |
) |
T5044 |
1997-1999 |
, |
denotes |
, |
T5045 |
1999-2006 |
VBG |
denotes |
leading |
T5046 |
2007-2009 |
IN |
denotes |
to |
T5047 |
2010-2017 |
NN |
denotes |
plasmid |
T5048 |
2023-2026 |
NN |
denotes |
DTA |
T5049 |
2018-2019 |
CD |
denotes |
3 |
T5050 |
2019-2020 |
SYM |
denotes |
′ |
T5051 |
2021-2022 |
SYM |
denotes |
+ |
T5052 |
2027-2030 |
CC |
denotes |
and |
T5053 |
2031-2032 |
-LRB- |
denotes |
( |
T5054 |
2032-2034 |
LS |
denotes |
ii |
T5055 |
2171-2178 |
VBD |
denotes |
deleted |
T5056 |
2034-2035 |
-RRB- |
denotes |
) |
T5057 |
2036-2043 |
IN |
denotes |
because |
T5058 |
2086-2094 |
VBZ |
denotes |
contains |
T5059 |
2044-2047 |
DT |
denotes |
the |
T5060 |
2061-2067 |
NN |
denotes |
region |
T5061 |
2048-2054 |
NN |
denotes |
Bsu36I |
T5062 |
2055-2060 |
NN |
denotes |
EcoRI |
T5063 |
2054-2055 |
HYPH |
denotes |
- |
T5064 |
2068-2078 |
RB |
denotes |
downstream |
T5065 |
2079-2081 |
IN |
denotes |
of |
T5066 |
2082-2085 |
NN |
denotes |
p53 |
T5067 |
2095-2105 |
JJ |
denotes |
repetitive |
T5068 |
2106-2115 |
NNS |
denotes |
sequences |
T5069 |
2116-2117 |
-LRB- |
denotes |
( |
T5070 |
2117-2119 |
NNP |
denotes |
F. |
T5071 |
2120-2126 |
NNP |
denotes |
Toledo |
T5072 |
2127-2130 |
CC |
denotes |
and |
T5073 |
2131-2133 |
NNP |
denotes |
G. |
T5074 |
2134-2136 |
NNP |
denotes |
M. |
T5075 |
2137-2141 |
NNP |
denotes |
Wahl |
T5076 |
2141-2143 |
, |
denotes |
, |
T5077 |
2143-2154 |
JJ |
denotes |
unpublished |
T5078 |
2155-2159 |
NNS |
denotes |
data |
T5079 |
2159-2160 |
-RRB- |
denotes |
) |
T5080 |
2160-2162 |
, |
denotes |
, |
T5081 |
2162-2164 |
PRP |
denotes |
we |
T5082 |
2165-2170 |
RB |
denotes |
later |
T5083 |
2179-2183 |
DT |
denotes |
this |
T5084 |
2184-2190 |
NN |
denotes |
region |
T5085 |
2190-2192 |
, |
denotes |
, |
T5086 |
2192-2194 |
TO |
denotes |
to |
T5087 |
2195-2201 |
VB |
denotes |
obtain |
T5088 |
2202-2209 |
NN |
denotes |
plasmid |
T5089 |
2215-2218 |
NN |
denotes |
DTA |
T5090 |
2210-2211 |
CD |
denotes |
3 |
T5091 |
2211-2212 |
SYM |
denotes |
′ |
T5092 |
2213-2214 |
SYM |
denotes |
+ |
T5093 |
2218-2219 |
. |
denotes |
. |
T5094 |
2219-2342 |
sentence |
denotes |
The 5′ homology consists of a 3.4 kb-long BamHI-PmlI fragment from intron 1 of p53 cloned in a modified pBS (plasmid p5′). |
T5095 |
2220-2223 |
DT |
denotes |
The |
T5096 |
2227-2235 |
NN |
denotes |
homology |
T5097 |
2224-2225 |
CD |
denotes |
5 |
T5098 |
2225-2226 |
SYM |
denotes |
′ |
T5099 |
2236-2244 |
VBZ |
denotes |
consists |
T5100 |
2245-2247 |
IN |
denotes |
of |
T5101 |
2248-2249 |
DT |
denotes |
a |
T5102 |
2273-2281 |
NN |
denotes |
fragment |
T5103 |
2250-2253 |
CD |
denotes |
3.4 |
T5104 |
2254-2256 |
NN |
denotes |
kb |
T5105 |
2257-2261 |
JJ |
denotes |
long |
T5106 |
2256-2257 |
HYPH |
denotes |
- |
T5107 |
2262-2267 |
NN |
denotes |
BamHI |
T5108 |
2268-2272 |
NN |
denotes |
PmlI |
T5109 |
2267-2268 |
HYPH |
denotes |
- |
T5110 |
2282-2286 |
IN |
denotes |
from |
T5111 |
2287-2293 |
NN |
denotes |
intron |
T5112 |
2294-2295 |
CD |
denotes |
1 |
T5113 |
2296-2298 |
IN |
denotes |
of |
T5114 |
2299-2302 |
NN |
denotes |
p53 |
T5115 |
2303-2309 |
VBN |
denotes |
cloned |
T5116 |
2310-2312 |
IN |
denotes |
in |
T5117 |
2313-2314 |
DT |
denotes |
a |
T5118 |
2324-2327 |
NN |
denotes |
pBS |
T5119 |
2315-2323 |
VBN |
denotes |
modified |
T5120 |
2328-2329 |
-LRB- |
denotes |
( |
T5121 |
2329-2336 |
NN |
denotes |
plasmid |
T5122 |
2337-2339 |
NN |
denotes |
p5 |
T5123 |
2339-2340 |
SYM |
denotes |
′ |
T5124 |
2340-2341 |
-RRB- |
denotes |
) |
T5125 |
2341-2342 |
. |
denotes |
. |
T5126 |
2342-2483 |
sentence |
denotes |
Finally, appropriate fragments from plasmids p5′, L3-p53PmlEagPuroΔTK-1L, and 3′ + DTA were assembled in a modified pSP72 plasmid (Promega). |
T5127 |
2343-2350 |
RB |
denotes |
Finally |
T5128 |
2435-2444 |
VBN |
denotes |
assembled |
T5129 |
2350-2352 |
, |
denotes |
, |
T5130 |
2352-2363 |
JJ |
denotes |
appropriate |
T5131 |
2364-2373 |
NNS |
denotes |
fragments |
T5132 |
2374-2378 |
IN |
denotes |
from |
T5133 |
2379-2387 |
NNS |
denotes |
plasmids |
T5134 |
2388-2390 |
NN |
denotes |
p5 |
T5135 |
2390-2391 |
SYM |
denotes |
′ |
T5136 |
2391-2393 |
, |
denotes |
, |
T5137 |
2393-2395 |
NN |
denotes |
L3 |
T5138 |
2413-2415 |
NN |
denotes |
1L |
T5139 |
2395-2396 |
HYPH |
denotes |
- |
T5140 |
2396-2412 |
NN |
denotes |
p53PmlEagPuroΔTK |
T5141 |
2412-2413 |
HYPH |
denotes |
- |
T5142 |
2415-2417 |
, |
denotes |
, |
T5143 |
2417-2420 |
CC |
denotes |
and |
T5144 |
2421-2422 |
CD |
denotes |
3 |
T5145 |
2426-2429 |
NN |
denotes |
DTA |
T5146 |
2422-2423 |
SYM |
denotes |
′ |
T5147 |
2424-2425 |
SYM |
denotes |
+ |
T5148 |
2430-2434 |
VBD |
denotes |
were |
T5149 |
2445-2447 |
IN |
denotes |
in |
T5150 |
2448-2449 |
DT |
denotes |
a |
T5151 |
2465-2472 |
NN |
denotes |
plasmid |
T5152 |
2450-2458 |
VBN |
denotes |
modified |
T5153 |
2459-2464 |
NN |
denotes |
pSP72 |
T5154 |
2473-2474 |
-LRB- |
denotes |
( |
T5155 |
2474-2481 |
NNP |
denotes |
Promega |
T5156 |
2481-2482 |
-RRB- |
denotes |
) |
T5157 |
2482-2483 |
. |
denotes |
. |
T5158 |
2483-2729 |
sentence |
denotes |
Plasmid Flox, the resulting targeting construct, was verified by restriction analysis, then sequenced using 30 primers chosen to precisely verify all p53 coding sequences, all exon–intron junctions and the sequences at and around the loxP sites. |
T5159 |
2484-2491 |
NN |
denotes |
Plasmid |
T5160 |
2492-2496 |
NN |
denotes |
Flox |
T5161 |
2537-2545 |
VBN |
denotes |
verified |
T5162 |
2496-2498 |
, |
denotes |
, |
T5163 |
2498-2501 |
DT |
denotes |
the |
T5164 |
2522-2531 |
NN |
denotes |
construct |
T5165 |
2502-2511 |
VBG |
denotes |
resulting |
T5166 |
2512-2521 |
NN |
denotes |
targeting |
T5167 |
2531-2533 |
, |
denotes |
, |
T5168 |
2533-2536 |
VBD |
denotes |
was |
T5169 |
2546-2548 |
IN |
denotes |
by |
T5170 |
2549-2560 |
NN |
denotes |
restriction |
T5171 |
2561-2569 |
NN |
denotes |
analysis |
T5172 |
2569-2571 |
, |
denotes |
, |
T5173 |
2571-2575 |
RB |
denotes |
then |
T5174 |
2576-2585 |
VBN |
denotes |
sequenced |
T5175 |
2586-2591 |
VBG |
denotes |
using |
T5176 |
2592-2594 |
CD |
denotes |
30 |
T5177 |
2595-2602 |
NNS |
denotes |
primers |
T5178 |
2603-2609 |
VBN |
denotes |
chosen |
T5179 |
2610-2612 |
TO |
denotes |
to |
T5180 |
2623-2629 |
VB |
denotes |
verify |
T5181 |
2613-2622 |
RB |
denotes |
precisely |
T5182 |
2630-2633 |
DT |
denotes |
all |
T5183 |
2645-2654 |
NNS |
denotes |
sequences |
T5184 |
2634-2637 |
NN |
denotes |
p53 |
T5185 |
2638-2644 |
NN |
denotes |
coding |
T5186 |
2654-2656 |
, |
denotes |
, |
T5187 |
2656-2659 |
DT |
denotes |
all |
T5188 |
2672-2681 |
NNS |
denotes |
junctions |
T5189 |
2660-2664 |
NN |
denotes |
exon |
T5190 |
2665-2671 |
NN |
denotes |
intron |
T5191 |
2664-2665 |
HYPH |
denotes |
– |
T5192 |
2682-2685 |
CC |
denotes |
and |
T5193 |
2686-2689 |
DT |
denotes |
the |
T5194 |
2690-2699 |
NNS |
denotes |
sequences |
T5195 |
2700-2702 |
IN |
denotes |
at |
T5196 |
2703-2706 |
CC |
denotes |
and |
T5197 |
2707-2713 |
IN |
denotes |
around |
T5198 |
2714-2717 |
DT |
denotes |
the |
T5199 |
2723-2728 |
NNS |
denotes |
sites |
T5200 |
2718-2722 |
NN |
denotes |
loxP |
T5201 |
2728-2729 |
. |
denotes |
. |
R1050 |
T4611 |
T4612 |
compound |
Targeting,construct |
R1051 |
T4613 |
T4612 |
prep |
for,construct |
R1052 |
T4614 |
T4615 |
det |
a,locus |
R1053 |
T4615 |
T4613 |
pobj |
locus,for |
R1054 |
T4616 |
T4615 |
nmod |
p53,locus |
R1055 |
T4617 |
T4618 |
npadvmod |
RMCE,ready |
R1056 |
T4618 |
T4615 |
amod |
ready,locus |
R1057 |
T4619 |
T4618 |
punct |
-,ready |
R1058 |
T4621 |
T4622 |
nsubj |
Details,are |
R1059 |
T4623 |
T4621 |
prep |
for,Details |
R1060 |
T4624 |
T4625 |
compound |
plasmid,construction |
R1061 |
T4625 |
T4623 |
pobj |
construction,for |
R1062 |
T4626 |
T4622 |
acomp |
available,are |
R1063 |
T4627 |
T4622 |
prep |
upon,are |
R1064 |
T4628 |
T4627 |
pobj |
request,upon |
R1065 |
T4629 |
T4622 |
prep |
owing,are |
R1066 |
T4630 |
T4629 |
prep |
to,owing |
R1067 |
T4631 |
T4632 |
compound |
space,limitations |
R1068 |
T4632 |
T4630 |
pobj |
limitations,to |
R1069 |
T4633 |
T4632 |
acl |
mandating,limitations |
R1070 |
T4634 |
T4635 |
det |
a,outline |
R1071 |
T4635 |
T4633 |
dobj |
outline,mandating |
R1072 |
T4636 |
T4635 |
amod |
brief,outline |
R1073 |
T4637 |
T4622 |
punct |
.,are |
R1074 |
T4639 |
T4640 |
nsubj |
We,started |
R1075 |
T4641 |
T4640 |
prep |
from,started |
R1076 |
T4642 |
T4643 |
det |
a,1L |
R1077 |
T4643 |
T4641 |
pobj |
1L,from |
R1078 |
T4644 |
T4643 |
compound |
plasmid,1L |
R1079 |
T4645 |
T4643 |
compound |
L3,1L |
R1080 |
T4646 |
T4643 |
punct |
-,1L |
R1081 |
T4647 |
T4643 |
acl |
containing,1L |
R1082 |
T4648 |
T4649 |
amod |
heterologous,sites |
R1083 |
T4649 |
T4647 |
dobj |
sites,containing |
R1084 |
T4650 |
T4649 |
compound |
loxP,sites |
R1085 |
T4651 |
T4652 |
punct |
(,is |
R1086 |
T4652 |
T4640 |
parataxis |
is,started |
R1087 |
T4653 |
T4654 |
nsubj |
L3,is |
R1088 |
T4654 |
T4652 |
ccomp |
is,is |
R1089 |
T4655 |
T4656 |
det |
the,loxP257 |
R1090 |
T4656 |
T4654 |
attr |
loxP257,is |
R1091 |
T4657 |
T4656 |
amod |
mutant,loxP257 |
R1092 |
T4658 |
T4659 |
advmod |
recently,described |
R1093 |
T4659 |
T4656 |
acl |
described,loxP257 |
R1094 |
T4660 |
T4661 |
punct |
(,14 |
R1095 |
T4661 |
T4654 |
parataxis |
14,is |
R1096 |
T4662 |
T4661 |
punct |
),14 |
R1097 |
T4663 |
T4652 |
punct |
", ",is |
R1098 |
T4664 |
T4652 |
nsubj |
1L,is |
R1099 |
T4665 |
T4666 |
det |
an,loxP |
R1100 |
T4666 |
T4652 |
attr |
loxP,is |
R1101 |
T4667 |
T4666 |
amod |
inverted,loxP |
R1102 |
T4668 |
T4666 |
compound |
WT,loxP |
R1103 |
T4669 |
T4652 |
punct |
),is |
R1104 |
T4670 |
T4640 |
punct |
.,started |
R1105 |
T4672 |
T4673 |
det |
The,loxP |
R1106 |
T4673 |
T4675 |
nsubj |
loxP,differ |
R1107 |
T4674 |
T4673 |
compound |
WT,loxP |
R1108 |
T4675 |
T4678 |
ccomp |
differ,is |
R1109 |
T4676 |
T4673 |
cc |
and,loxP |
R1110 |
T4677 |
T4673 |
conj |
loxP257,loxP |
R1111 |
T4679 |
T4675 |
prep |
in,differ |
R1112 |
T4680 |
T4681 |
poss |
their,sequences |
R1113 |
T4681 |
T4679 |
pobj |
sequences,in |
R1114 |
T4682 |
T4681 |
compound |
spacer,sequences |
R1115 |
T4683 |
T4678 |
punct |
: ,is |
R1116 |
T4684 |
T4685 |
det |
the,sequence |
R1117 |
T4685 |
T4678 |
nsubj |
sequence,is |
R1118 |
T4686 |
T4685 |
compound |
spacer,sequence |
R1119 |
T4687 |
T4678 |
attr |
5,is |
R1120 |
T4688 |
T4687 |
punct |
′,5 |
R1121 |
T4689 |
T4687 |
punct |
-,5 |
R1122 |
T4690 |
T4687 |
amod |
ATGTATGC,5 |
R1123 |
T4691 |
T4687 |
punct |
-,5 |
R1124 |
T4692 |
T4687 |
nummod |
3,5 |
R1125 |
T4693 |
T4687 |
punct |
′,5 |
R1126 |
T4694 |
T4687 |
prep |
for,5 |
R1127 |
T4695 |
T4696 |
compound |
WT,loxP |
R1128 |
T4696 |
T4694 |
pobj |
loxP,for |
R1129 |
T4697 |
T4687 |
cc |
and,5 |
R1130 |
T4698 |
T4699 |
nummod |
5,AAGTCTCC |
R1131 |
T4699 |
T4687 |
conj |
AAGTCTCC,5 |
R1132 |
T4700 |
T4698 |
punct |
′,5 |
R1133 |
T4701 |
T4699 |
punct |
-,AAGTCTCC |
R1134 |
T4702 |
T4699 |
punct |
-,AAGTCTCC |
R1135 |
T4703 |
T4699 |
nummod |
3,AAGTCTCC |
R1136 |
T4704 |
T4699 |
punct |
′,AAGTCTCC |
R1137 |
T4705 |
T4699 |
prep |
for,AAGTCTCC |
R1138 |
T4706 |
T4705 |
pobj |
loxP257,for |
R1139 |
T4707 |
T4678 |
punct |
.,is |
R1140 |
T4709 |
T4710 |
det |
The,mutations |
R1141 |
T4710 |
T4712 |
nsubj |
mutations,prevent |
R1142 |
T4711 |
T4710 |
nummod |
three,mutations |
R1143 |
T4712 |
T4718 |
ccomp |
prevent,occurred |
R1144 |
T4713 |
T4710 |
prep |
in,mutations |
R1145 |
T4714 |
T4715 |
det |
the,sequence |
R1146 |
T4715 |
T4713 |
pobj |
sequence,in |
R1147 |
T4716 |
T4715 |
compound |
loxP257,sequence |
R1148 |
T4717 |
T4715 |
compound |
spacer,sequence |
R1149 |
T4719 |
T4720 |
nsubj |
it,recombine |
R1150 |
T4720 |
T4712 |
ccomp |
recombine,prevent |
R1151 |
T4721 |
T4720 |
aux |
to,recombine |
R1152 |
T4722 |
T4720 |
prep |
with,recombine |
R1153 |
T4723 |
T4724 |
compound |
WT,loxP |
R1154 |
T4724 |
T4722 |
pobj |
loxP,with |
R1155 |
T4725 |
T4712 |
punct |
", ",prevent |
R1156 |
T4726 |
T4712 |
advcl |
ensuring,prevent |
R1157 |
T4727 |
T4728 |
amod |
efficient,RMCE |
R1158 |
T4728 |
T4726 |
dobj |
RMCE,ensuring |
R1159 |
T4729 |
T4726 |
prep |
in,ensuring |
R1160 |
T4730 |
T4731 |
amod |
several,lines |
R1161 |
T4731 |
T4729 |
pobj |
lines,in |
R1162 |
T4732 |
T4731 |
compound |
cell,lines |
R1163 |
T4733 |
T4718 |
punct |
: ,occurred |
R1164 |
T4734 |
T4735 |
amod |
accurate,RMCE |
R1165 |
T4735 |
T4718 |
nsubj |
RMCE,occurred |
R1166 |
T4736 |
T4735 |
prep |
with,RMCE |
R1167 |
T4737 |
T4738 |
det |
these,sites |
R1168 |
T4738 |
T4736 |
pobj |
sites,with |
R1169 |
T4739 |
T4738 |
compound |
loxP,sites |
R1170 |
T4740 |
T4718 |
prep |
with,occurred |
R1171 |
T4741 |
T4742 |
det |
an,frequency |
R1172 |
T4742 |
T4740 |
pobj |
frequency,with |
R1173 |
T4743 |
T4742 |
amod |
average,frequency |
R1174 |
T4744 |
T4742 |
prep |
of,frequency |
R1175 |
T4745 |
T4746 |
nummod |
81,% |
R1176 |
T4746 |
T4744 |
pobj |
%,of |
R1177 |
T4747 |
T4742 |
prep |
at,frequency |
R1178 |
T4748 |
T4749 |
nummod |
two,loci |
R1179 |
T4749 |
T4747 |
pobj |
loci,at |
R1180 |
T4750 |
T4749 |
prep |
in,loci |
R1181 |
T4751 |
T4752 |
compound |
CHO,cells |
R1182 |
T4752 |
T4750 |
pobj |
cells,in |
R1183 |
T4753 |
T4742 |
cc |
and,frequency |
R1184 |
T4754 |
T4755 |
det |
an,frequency |
R1185 |
T4755 |
T4742 |
conj |
frequency,frequency |
R1186 |
T4756 |
T4755 |
amod |
average,frequency |
R1187 |
T4757 |
T4755 |
prep |
of,frequency |
R1188 |
T4758 |
T4759 |
nummod |
69,% |
R1189 |
T4759 |
T4757 |
pobj |
%,of |
R1190 |
T4760 |
T4755 |
prep |
at,frequency |
R1191 |
T4761 |
T4762 |
nummod |
four,loci |
R1192 |
T4762 |
T4760 |
pobj |
loci,at |
R1193 |
T4763 |
T4762 |
prep |
in,loci |
R1194 |
T4764 |
T4765 |
compound |
Hela,cells |
R1195 |
T4765 |
T4763 |
pobj |
cells,in |
R1196 |
T4766 |
T4767 |
punct |
(,14 |
R1197 |
T4767 |
T4718 |
parataxis |
14,occurred |
R1198 |
T4768 |
T4767 |
punct |
),14 |
R1199 |
T4769 |
T4718 |
punct |
.,occurred |
R1200 |
T4771 |
T4772 |
det |
The,plasmid |
R1201 |
T4772 |
T4776 |
nsubjpass |
plasmid,modified |
R1202 |
T4773 |
T4774 |
compound |
L3,1L |
R1203 |
T4774 |
T4772 |
compound |
1L,plasmid |
R1204 |
T4775 |
T4774 |
punct |
-,1L |
R1205 |
T4777 |
T4776 |
auxpass |
was,modified |
R1206 |
T4778 |
T4776 |
advmod |
first,modified |
R1207 |
T4779 |
T4780 |
aux |
to,include |
R1208 |
T4780 |
T4776 |
advcl |
include,modified |
R1209 |
T4781 |
T4782 |
det |
a,ClaI |
R1210 |
T4782 |
T4780 |
dobj |
ClaI,include |
R1211 |
T4783 |
T4782 |
cc |
and,ClaI |
R1212 |
T4784 |
T4785 |
det |
a,site |
R1213 |
T4785 |
T4782 |
conj |
site,ClaI |
R1214 |
T4786 |
T4785 |
compound |
FseI,site |
R1215 |
T4787 |
T4780 |
prep |
between,include |
R1216 |
T4788 |
T4789 |
det |
the,sites |
R1217 |
T4789 |
T4787 |
pobj |
sites,between |
R1218 |
T4790 |
T4789 |
compound |
LoxP,sites |
R1219 |
T4791 |
T4776 |
punct |
", ",modified |
R1220 |
T4792 |
T4776 |
advcl |
leading,modified |
R1221 |
T4793 |
T4792 |
prep |
to,leading |
R1222 |
T4794 |
T4795 |
compound |
plasmid,1L |
R1223 |
T4795 |
T4793 |
pobj |
1L,to |
R1224 |
T4796 |
T4795 |
compound |
L3,1L |
R1225 |
T4797 |
T4795 |
punct |
-,1L |
R1226 |
T4798 |
T4795 |
compound |
CF,1L |
R1227 |
T4799 |
T4795 |
punct |
-,1L |
R1228 |
T4800 |
T4776 |
punct |
.,modified |
R1229 |
T4802 |
T4803 |
nsubj |
We,modified |
R1230 |
T4804 |
T4803 |
advmod |
next,modified |
R1231 |
T4805 |
T4806 |
det |
a,plasmid |
R1232 |
T4806 |
T4803 |
dobj |
plasmid,modified |
R1233 |
T4807 |
T4806 |
compound |
puroΔTK,plasmid |
R1234 |
T4808 |
T4806 |
punct |
(,plasmid |
R1235 |
T4809 |
T4806 |
appos |
YTC37,plasmid |
R1236 |
T4810 |
T4809 |
punct |
", ",YTC37 |
R1237 |
T4811 |
T4812 |
det |
a,gift |
R1238 |
T4812 |
T4809 |
appos |
gift,YTC37 |
R1239 |
T4813 |
T4812 |
compound |
kind,gift |
R1240 |
T4814 |
T4812 |
prep |
from,gift |
R1241 |
T4815 |
T4816 |
compound |
A.,Bradley |
R1242 |
T4816 |
T4814 |
pobj |
Bradley,from |
R1243 |
T4817 |
T4803 |
punct |
),modified |
R1244 |
T4818 |
T4803 |
prep |
by,modified |
R1245 |
T4819 |
T4818 |
pcomp |
using,by |
R1246 |
T4820 |
T4819 |
dobj |
oligonucleotides,using |
R1247 |
T4821 |
T4822 |
aux |
to,destroy |
R1248 |
T4822 |
T4819 |
advcl |
destroy,using |
R1249 |
T4823 |
T4824 |
det |
a,site |
R1250 |
T4824 |
T4822 |
dobj |
site,destroy |
R1251 |
T4825 |
T4824 |
compound |
NotI,site |
R1252 |
T4826 |
T4824 |
advmod |
downstream,site |
R1253 |
T4827 |
T4826 |
prep |
of,downstream |
R1254 |
T4828 |
T4829 |
det |
the,gene |
R1255 |
T4829 |
T4827 |
pobj |
gene,of |
R1256 |
T4830 |
T4829 |
compound |
puroΔTK,gene |
R1257 |
T4831 |
T4822 |
cc |
and,destroy |
R1258 |
T4832 |
T4822 |
conj |
introduce,destroy |
R1259 |
T4833 |
T4834 |
det |
a,site |
R1260 |
T4834 |
T4832 |
dobj |
site,introduce |
R1261 |
T4835 |
T4834 |
compound |
NotI,site |
R1262 |
T4836 |
T4832 |
advmod |
upstream,introduce |
R1263 |
T4837 |
T4832 |
punct |
", ",introduce |
R1264 |
T4838 |
T4832 |
cc |
and,introduce |
R1265 |
T4839 |
T4840 |
det |
a,site |
R1266 |
T4840 |
T4832 |
conj |
site,introduce |
R1267 |
T4841 |
T4840 |
compound |
FseI,site |
R1268 |
T4842 |
T4840 |
advmod |
downstream,site |
R1269 |
T4843 |
T4842 |
prep |
of,downstream |
R1270 |
T4844 |
T4845 |
det |
the,gene |
R1271 |
T4845 |
T4843 |
pobj |
gene,of |
R1272 |
T4846 |
T4847 |
punct |
(,leading |
R1273 |
T4847 |
T4840 |
parataxis |
leading,site |
R1274 |
T4848 |
T4847 |
prep |
to,leading |
R1275 |
T4849 |
T4850 |
compound |
plasmid,F |
R1276 |
T4850 |
T4848 |
pobj |
F,to |
R1277 |
T4851 |
T4850 |
compound |
CN,F |
R1278 |
T4852 |
T4850 |
punct |
-,F |
R1279 |
T4853 |
T4850 |
compound |
PuroΔTK,F |
R1280 |
T4854 |
T4850 |
punct |
-,F |
R1281 |
T4855 |
T4847 |
punct |
),leading |
R1282 |
T4856 |
T4803 |
punct |
.,modified |
R1283 |
T4858 |
T4859 |
advmod |
Next,subcloned |
R1284 |
T4860 |
T4859 |
punct |
", ",subcloned |
R1285 |
T4861 |
T4862 |
det |
a,fragment |
R1286 |
T4862 |
T4859 |
nsubjpass |
fragment,subcloned |
R1287 |
T4863 |
T4864 |
nmod |
PmlI,MfeI |
R1288 |
T4864 |
T4862 |
nmod |
MfeI,fragment |
R1289 |
T4865 |
T4864 |
punct |
-,MfeI |
R1290 |
T4866 |
T4867 |
nummod |
6.3,kb |
R1291 |
T4867 |
T4862 |
compound |
kb,fragment |
R1292 |
T4868 |
T4862 |
prep |
from,fragment |
R1293 |
T4869 |
T4868 |
pobj |
Trp53,from |
R1294 |
T4870 |
T4859 |
auxpass |
was,subcloned |
R1295 |
T4871 |
T4859 |
prep |
in,subcloned |
R1296 |
T4872 |
T4873 |
det |
a,KSII+ |
R1297 |
T4873 |
T4871 |
pobj |
KSII+,in |
R1298 |
T4874 |
T4873 |
amod |
modified,KSII+ |
R1299 |
T4875 |
T4873 |
compound |
pBluescript,KSII+ |
R1300 |
T4876 |
T4877 |
punct |
(,pBS |
R1301 |
T4877 |
T4873 |
parataxis |
pBS,KSII+ |
R1302 |
T4878 |
T4877 |
punct |
", ",pBS |
R1303 |
T4879 |
T4877 |
npadvmod |
Stratagene,pBS |
R1304 |
T4880 |
T4877 |
punct |
),pBS |
R1305 |
T4881 |
T4859 |
punct |
", ",subcloned |
R1306 |
T4882 |
T4859 |
cc |
and,subcloned |
R1307 |
T4883 |
T4884 |
det |
the,plasmid |
R1308 |
T4884 |
T4886 |
nsubjpass |
plasmid,digested |
R1309 |
T4885 |
T4884 |
amod |
resulting,plasmid |
R1310 |
T4886 |
T4859 |
conj |
digested,subcloned |
R1311 |
T4887 |
T4886 |
auxpass |
was,digested |
R1312 |
T4888 |
T4886 |
prep |
with,digested |
R1313 |
T4889 |
T4888 |
pobj |
SwaI,with |
R1314 |
T4890 |
T4891 |
aux |
to,introduce |
R1315 |
T4891 |
T4886 |
advcl |
introduce,digested |
R1316 |
T4892 |
T4893 |
det |
an,site |
R1317 |
T4893 |
T4891 |
dobj |
site,introduce |
R1318 |
T4894 |
T4893 |
compound |
EagI,site |
R1319 |
T4895 |
T4891 |
punct |
", ",introduce |
R1320 |
T4896 |
T4891 |
advcl |
leading,introduce |
R1321 |
T4897 |
T4896 |
prep |
to,leading |
R1322 |
T4898 |
T4897 |
pobj |
p53PmlEag,to |
R1323 |
T4899 |
T4898 |
punct |
", ",p53PmlEag |
R1324 |
T4900 |
T4901 |
det |
a,plasmid |
R1325 |
T4901 |
T4898 |
appos |
plasmid,p53PmlEag |
R1326 |
T4902 |
T4901 |
acl |
containing,plasmid |
R1327 |
T4903 |
T4904 |
nmod |
exons,11 |
R1328 |
T4904 |
T4902 |
dobj |
11,containing |
R1329 |
T4905 |
T4904 |
nummod |
2,11 |
R1330 |
T4906 |
T4904 |
punct |
–,11 |
R1331 |
T4907 |
T4904 |
prep |
of,11 |
R1332 |
T4908 |
T4907 |
pobj |
p53,of |
R1333 |
T4909 |
T4886 |
punct |
.,digested |
R1334 |
T4911 |
T4912 |
nsubj |
We,inserted |
R1335 |
T4913 |
T4912 |
advmod |
then,inserted |
R1336 |
T4914 |
T4915 |
det |
a,fragment |
R1337 |
T4915 |
T4912 |
dobj |
fragment,inserted |
R1338 |
T4916 |
T4917 |
nummod |
5.5,kb |
R1339 |
T4917 |
T4915 |
compound |
kb,fragment |
R1340 |
T4918 |
T4919 |
compound |
ClaI,EagI |
R1341 |
T4919 |
T4915 |
compound |
EagI,fragment |
R1342 |
T4920 |
T4919 |
punct |
-,EagI |
R1343 |
T4921 |
T4915 |
prep |
from,fragment |
R1344 |
T4922 |
T4921 |
pobj |
p53PmlEag,from |
R1345 |
T4923 |
T4915 |
prep |
in,fragment |
R1346 |
T4924 |
T4925 |
compound |
plasmid,F |
R1347 |
T4925 |
T4923 |
pobj |
F,in |
R1348 |
T4926 |
T4925 |
compound |
CN,F |
R1349 |
T4927 |
T4925 |
punct |
-,F |
R1350 |
T4928 |
T4925 |
compound |
PuroΔTK,F |
R1351 |
T4929 |
T4925 |
punct |
-,F |
R1352 |
T4930 |
T4915 |
acl |
digested,fragment |
R1353 |
T4931 |
T4930 |
agent |
by,digested |
R1354 |
T4932 |
T4931 |
pobj |
ClaI,by |
R1355 |
T4933 |
T4932 |
cc |
and,ClaI |
R1356 |
T4934 |
T4932 |
conj |
NotI,ClaI |
R1357 |
T4935 |
T4912 |
punct |
", ",inserted |
R1358 |
T4936 |
T4912 |
cc |
and,inserted |
R1359 |
T4937 |
T4912 |
conj |
inserted,inserted |
R1360 |
T4938 |
T4939 |
det |
the,fragment |
R1361 |
T4939 |
T4937 |
dobj |
fragment,inserted |
R1362 |
T4940 |
T4939 |
amod |
resulting,fragment |
R1363 |
T4941 |
T4937 |
prep |
between,inserted |
R1364 |
T4942 |
T4943 |
det |
the,sites |
R1365 |
T4943 |
T4941 |
pobj |
sites,between |
R1366 |
T4944 |
T4943 |
compound |
loxP,sites |
R1367 |
T4945 |
T4943 |
prep |
of,sites |
R1368 |
T4946 |
T4947 |
compound |
L3,1L |
R1369 |
T4947 |
T4945 |
pobj |
1L,of |
R1370 |
T4948 |
T4947 |
punct |
-,1L |
R1371 |
T4949 |
T4947 |
compound |
CF,1L |
R1372 |
T4950 |
T4947 |
punct |
-,1L |
R1373 |
T4951 |
T4937 |
prep |
by,inserted |
R1374 |
T4952 |
T4953 |
nmod |
ClaI,digestion |
R1375 |
T4953 |
T4951 |
pobj |
digestion,by |
R1376 |
T4954 |
T4952 |
cc |
and,ClaI |
R1377 |
T4955 |
T4952 |
conj |
FseI,ClaI |
R1378 |
T4956 |
T4937 |
punct |
", ",inserted |
R1379 |
T4957 |
T4937 |
advcl |
leading,inserted |
R1380 |
T4958 |
T4957 |
prep |
to,leading |
R1381 |
T4959 |
T4960 |
compound |
L3,1L |
R1382 |
T4960 |
T4958 |
pobj |
1L,to |
R1383 |
T4961 |
T4960 |
punct |
-,1L |
R1384 |
T4962 |
T4960 |
compound |
p53PmlEagPuroΔTK,1L |
R1385 |
T4963 |
T4960 |
punct |
-,1L |
R1386 |
T4964 |
T4912 |
punct |
.,inserted |
R1387 |
T4966 |
T4967 |
nsubj |
We,engineered |
R1388 |
T4967 |
T4969 |
ccomp |
engineered,performed |
R1389 |
T4968 |
T4967 |
advmod |
next,engineered |
R1390 |
T4970 |
T4971 |
det |
a,plasmid |
R1391 |
T4971 |
T4967 |
dobj |
plasmid,engineered |
R1392 |
T4972 |
T4971 |
acl |
containing,plasmid |
R1393 |
T4973 |
T4974 |
det |
the,region |
R1394 |
T4974 |
T4972 |
dobj |
region,containing |
R1395 |
T4975 |
T4974 |
prep |
for,region |
R1396 |
T4976 |
T4975 |
pobj |
3,for |
R1397 |
T4977 |
T4976 |
punct |
′,3 |
R1398 |
T4978 |
T4976 |
punct |
-,3 |
R1399 |
T4979 |
T4976 |
amod |
homology,3 |
R1400 |
T4980 |
T4972 |
advmod |
downstream,containing |
R1401 |
T4981 |
T4980 |
prep |
of,downstream |
R1402 |
T4982 |
T4983 |
det |
the,gene |
R1403 |
T4983 |
T4981 |
pobj |
gene,of |
R1404 |
T4984 |
T4983 |
compound |
p53,gene |
R1405 |
T4985 |
T4983 |
cc |
and,gene |
R1406 |
T4986 |
T4987 |
det |
the,gene |
R1407 |
T4987 |
T4983 |
conj |
gene,gene |
R1408 |
T4988 |
T4987 |
compound |
DTA,gene |
R1409 |
T4989 |
T4967 |
prep |
in,engineered |
R1410 |
T4990 |
T4991 |
nummod |
two,steps |
R1411 |
T4991 |
T4989 |
pobj |
steps,in |
R1412 |
T4992 |
T4969 |
punct |
: ,performed |
R1413 |
T4993 |
T4994 |
punct |
(,i |
R1414 |
T4994 |
T4969 |
meta |
i,performed |
R1415 |
T4995 |
T4994 |
punct |
),i |
R1416 |
T4996 |
T4969 |
nsubj |
we,performed |
R1417 |
T4997 |
T4998 |
det |
a,ligation |
R1418 |
T4998 |
T4969 |
dobj |
ligation,performed |
R1419 |
T4999 |
T5000 |
nummod |
three,way |
R1420 |
T5000 |
T4998 |
compound |
way,ligation |
R1421 |
T5001 |
T5000 |
punct |
-,way |
R1422 |
T5002 |
T4998 |
prep |
between,ligation |
R1423 |
T5003 |
T5004 |
det |
a,pBS |
R1424 |
T5004 |
T5002 |
pobj |
pBS,between |
R1425 |
T5005 |
T5004 |
amod |
modified,pBS |
R1426 |
T5006 |
T5004 |
acl |
digested,pBS |
R1427 |
T5007 |
T5006 |
agent |
by,digested |
R1428 |
T5008 |
T5007 |
pobj |
HindIII,by |
R1429 |
T5009 |
T5008 |
cc |
and,HindIII |
R1430 |
T5010 |
T5008 |
conj |
NotI,HindIII |
R1431 |
T5011 |
T5004 |
punct |
", ",pBS |
R1432 |
T5012 |
T5013 |
det |
a,fragment |
R1433 |
T5013 |
T5004 |
conj |
fragment,pBS |
R1434 |
T5014 |
T5015 |
compound |
HindIII,EcoRI |
R1435 |
T5015 |
T5013 |
compound |
EcoRI,fragment |
R1436 |
T5016 |
T5015 |
punct |
-,EcoRI |
R1437 |
T5017 |
T5013 |
prep |
from,fragment |
R1438 |
T5018 |
T5017 |
pobj |
Trp53,from |
R1439 |
T5019 |
T5013 |
prep |
for,fragment |
R1440 |
T5020 |
T5021 |
nummod |
3,homology |
R1441 |
T5021 |
T5019 |
pobj |
homology,for |
R1442 |
T5022 |
T5020 |
punct |
′,3 |
R1443 |
T5023 |
T5013 |
cc |
and,fragment |
R1444 |
T5024 |
T5025 |
det |
an,fragment |
R1445 |
T5025 |
T5013 |
conj |
fragment,fragment |
R1446 |
T5026 |
T5027 |
compound |
EcoRI,NotI |
R1447 |
T5027 |
T5025 |
compound |
NotI,fragment |
R1448 |
T5028 |
T5027 |
punct |
-,NotI |
R1449 |
T5029 |
T5025 |
acl |
containing,fragment |
R1450 |
T5030 |
T5031 |
det |
the,gene |
R1451 |
T5031 |
T5029 |
dobj |
gene,containing |
R1452 |
T5032 |
T5031 |
compound |
DTA,gene |
R1453 |
T5033 |
T5025 |
punct |
", ",fragment |
R1454 |
T5034 |
T5025 |
prep |
from,fragment |
R1455 |
T5035 |
T5036 |
compound |
plasmid,pgkdtabpa |
R1456 |
T5036 |
T5034 |
pobj |
pgkdtabpa,from |
R1457 |
T5037 |
T5025 |
punct |
(,fragment |
R1458 |
T5038 |
T5039 |
amod |
kind,gift |
R1459 |
T5039 |
T5025 |
appos |
gift,fragment |
R1460 |
T5040 |
T5039 |
prep |
of,gift |
R1461 |
T5041 |
T5042 |
compound |
P.,Soriano |
R1462 |
T5042 |
T5040 |
pobj |
Soriano,of |
R1463 |
T5043 |
T4969 |
punct |
),performed |
R1464 |
T5044 |
T4969 |
punct |
", ",performed |
R1465 |
T5045 |
T4969 |
advcl |
leading,performed |
R1466 |
T5046 |
T5045 |
prep |
to,leading |
R1467 |
T5047 |
T5048 |
nmod |
plasmid,DTA |
R1468 |
T5048 |
T5046 |
pobj |
DTA,to |
R1469 |
T5049 |
T5048 |
nummod |
3,DTA |
R1470 |
T5050 |
T5049 |
punct |
′,3 |
R1471 |
T5051 |
T5049 |
punct |
+,3 |
R1472 |
T5052 |
T4969 |
cc |
and,performed |
R1473 |
T5053 |
T5054 |
punct |
(,ii |
R1474 |
T5054 |
T5055 |
meta |
ii,deleted |
R1475 |
T5055 |
T4969 |
conj |
deleted,performed |
R1476 |
T5056 |
T5054 |
punct |
),ii |
R1477 |
T5057 |
T5058 |
mark |
because,contains |
R1478 |
T5058 |
T5055 |
advcl |
contains,deleted |
R1479 |
T5059 |
T5060 |
det |
the,region |
R1480 |
T5060 |
T5058 |
nsubj |
region,contains |
R1481 |
T5061 |
T5062 |
compound |
Bsu36I,EcoRI |
R1482 |
T5062 |
T5060 |
compound |
EcoRI,region |
R1483 |
T5063 |
T5062 |
punct |
-,EcoRI |
R1484 |
T5064 |
T5060 |
advmod |
downstream,region |
R1485 |
T5065 |
T5064 |
prep |
of,downstream |
R1486 |
T5066 |
T5065 |
pobj |
p53,of |
R1487 |
T5067 |
T5068 |
amod |
repetitive,sequences |
R1488 |
T5068 |
T5058 |
dobj |
sequences,contains |
R1489 |
T5069 |
T5070 |
punct |
(,F. |
R1490 |
T5070 |
T5058 |
meta |
F.,contains |
R1491 |
T5071 |
T5070 |
nmod |
Toledo,F. |
R1492 |
T5072 |
T5070 |
cc |
and,F. |
R1493 |
T5073 |
T5070 |
conj |
G.,F. |
R1494 |
T5074 |
T5073 |
nmod |
M.,G. |
R1495 |
T5075 |
T5073 |
nmod |
Wahl,G. |
R1496 |
T5076 |
T5073 |
punct |
", ",G. |
R1497 |
T5077 |
T5078 |
amod |
unpublished,data |
R1498 |
T5078 |
T5073 |
conj |
data,G. |
R1499 |
T5079 |
T5078 |
punct |
),data |
R1500 |
T5080 |
T5055 |
punct |
", ",deleted |
R1501 |
T5081 |
T5055 |
nsubj |
we,deleted |
R1502 |
T5082 |
T5055 |
advmod |
later,deleted |
R1503 |
T5083 |
T5084 |
det |
this,region |
R1504 |
T5084 |
T5055 |
dobj |
region,deleted |
R1505 |
T5085 |
T5055 |
punct |
", ",deleted |
R1506 |
T5086 |
T5087 |
aux |
to,obtain |
R1507 |
T5087 |
T5055 |
advcl |
obtain,deleted |
R1508 |
T5088 |
T5089 |
nmod |
plasmid,DTA |
R1509 |
T5089 |
T5087 |
dobj |
DTA,obtain |
R1510 |
T5090 |
T5089 |
nummod |
3,DTA |
R1511 |
T5091 |
T5090 |
punct |
′,3 |
R1512 |
T5092 |
T5090 |
punct |
+,3 |
R1513 |
T5093 |
T4969 |
punct |
.,performed |
R1514 |
T5095 |
T5096 |
det |
The,homology |
R1515 |
T5096 |
T5099 |
nsubj |
homology,consists |
R1516 |
T5097 |
T5096 |
nummod |
5,homology |
R1517 |
T5098 |
T5097 |
punct |
′,5 |
R1518 |
T5100 |
T5099 |
prep |
of,consists |
R1519 |
T5101 |
T5102 |
det |
a,fragment |
R1520 |
T5102 |
T5100 |
pobj |
fragment,of |
R1521 |
T5103 |
T5104 |
nummod |
3.4,kb |
R1522 |
T5104 |
T5105 |
npadvmod |
kb,long |
R1523 |
T5105 |
T5102 |
amod |
long,fragment |
R1524 |
T5106 |
T5105 |
punct |
-,long |
R1525 |
T5107 |
T5108 |
compound |
BamHI,PmlI |
R1526 |
T5108 |
T5102 |
compound |
PmlI,fragment |
R1527 |
T5109 |
T5108 |
punct |
-,PmlI |
R1528 |
T5110 |
T5102 |
prep |
from,fragment |
R1529 |
T5111 |
T5110 |
pobj |
intron,from |
R1530 |
T5112 |
T5111 |
nummod |
1,intron |
R1531 |
T5113 |
T5111 |
prep |
of,intron |
R1532 |
T5114 |
T5113 |
pobj |
p53,of |
R1533 |
T5115 |
T5102 |
acl |
cloned,fragment |
R1534 |
T5116 |
T5115 |
prep |
in,cloned |
R1535 |
T5117 |
T5118 |
det |
a,pBS |
R1536 |
T5118 |
T5116 |
pobj |
pBS,in |
R1537 |
T5119 |
T5118 |
amod |
modified,pBS |
R1538 |
T5120 |
T5118 |
punct |
(,pBS |
R1539 |
T5121 |
T5122 |
compound |
plasmid,p5 |
R1540 |
T5122 |
T5118 |
appos |
p5,pBS |
R1541 |
T5123 |
T5122 |
punct |
′,p5 |
R1542 |
T5124 |
T5099 |
punct |
),consists |
R1543 |
T5125 |
T5099 |
punct |
.,consists |
R1544 |
T5127 |
T5128 |
advmod |
Finally,assembled |
R1545 |
T5129 |
T5128 |
punct |
", ",assembled |
R1546 |
T5130 |
T5131 |
amod |
appropriate,fragments |
R1547 |
T5131 |
T5128 |
nsubjpass |
fragments,assembled |
R1548 |
T5132 |
T5131 |
prep |
from,fragments |
R1549 |
T5133 |
T5134 |
compound |
plasmids,p5 |
R1550 |
T5134 |
T5132 |
pobj |
p5,from |
R1551 |
T5135 |
T5134 |
punct |
′,p5 |
R1552 |
T5136 |
T5134 |
punct |
", ",p5 |
R1553 |
T5137 |
T5138 |
compound |
L3,1L |
R1554 |
T5138 |
T5134 |
conj |
1L,p5 |
R1555 |
T5139 |
T5138 |
punct |
-,1L |
R1556 |
T5140 |
T5138 |
compound |
p53PmlEagPuroΔTK,1L |
R1557 |
T5141 |
T5138 |
punct |
-,1L |
R1558 |
T5142 |
T5138 |
punct |
", ",1L |
R1559 |
T5143 |
T5138 |
cc |
and,1L |
R1560 |
T5144 |
T5145 |
nummod |
3,DTA |
R1561 |
T5145 |
T5138 |
conj |
DTA,1L |
R1562 |
T5146 |
T5144 |
punct |
′,3 |
R1563 |
T5147 |
T5144 |
punct |
+,3 |
R1564 |
T5148 |
T5128 |
auxpass |
were,assembled |
R1565 |
T5149 |
T5128 |
prep |
in,assembled |
R1566 |
T5150 |
T5151 |
det |
a,plasmid |
R1567 |
T5151 |
T5149 |
pobj |
plasmid,in |
R1568 |
T5152 |
T5151 |
amod |
modified,plasmid |
R1569 |
T5153 |
T5151 |
compound |
pSP72,plasmid |
R1570 |
T5154 |
T5155 |
punct |
(,Promega |
R1571 |
T5155 |
T5151 |
parataxis |
Promega,plasmid |
R1572 |
T5156 |
T5155 |
punct |
),Promega |
R1573 |
T5157 |
T5128 |
punct |
.,assembled |
R1574 |
T5159 |
T5160 |
compound |
Plasmid,Flox |
R1575 |
T5160 |
T5161 |
nsubjpass |
Flox,verified |
R1576 |
T5162 |
T5160 |
punct |
", ",Flox |
R1577 |
T5163 |
T5164 |
det |
the,construct |
R1578 |
T5164 |
T5160 |
appos |
construct,Flox |
R1579 |
T5165 |
T5164 |
amod |
resulting,construct |
R1580 |
T5166 |
T5164 |
compound |
targeting,construct |
R1581 |
T5167 |
T5161 |
punct |
", ",verified |
R1582 |
T5168 |
T5161 |
auxpass |
was,verified |
R1583 |
T5169 |
T5161 |
prep |
by,verified |
R1584 |
T5170 |
T5171 |
compound |
restriction,analysis |
R1585 |
T5171 |
T5169 |
pobj |
analysis,by |
R1586 |
T5172 |
T5161 |
punct |
", ",verified |
R1587 |
T5173 |
T5161 |
advmod |
then,verified |
R1588 |
T5174 |
T5161 |
dep |
sequenced,verified |
R1589 |
T5175 |
T5174 |
advcl |
using,sequenced |
R1590 |
T5176 |
T5177 |
nummod |
30,primers |
R1591 |
T5177 |
T5175 |
dobj |
primers,using |
R1592 |
T5178 |
T5177 |
acl |
chosen,primers |
R1593 |
T5179 |
T5180 |
aux |
to,verify |
R1594 |
T5180 |
T5178 |
advcl |
verify,chosen |
R1595 |
T5181 |
T5180 |
advmod |
precisely,verify |
R1596 |
T5182 |
T5183 |
det |
all,sequences |
R1597 |
T5183 |
T5180 |
dobj |
sequences,verify |
R1598 |
T5184 |
T5185 |
compound |
p53,coding |
R1599 |
T5185 |
T5183 |
compound |
coding,sequences |
R1600 |
T5186 |
T5183 |
punct |
", ",sequences |
R1601 |
T5187 |
T5188 |
det |
all,junctions |
R1602 |
T5188 |
T5183 |
conj |
junctions,sequences |
R1603 |
T5189 |
T5190 |
compound |
exon,intron |
R1604 |
T5190 |
T5188 |
compound |
intron,junctions |
R1605 |
T5191 |
T5190 |
punct |
–,intron |
R1606 |
T5192 |
T5188 |
cc |
and,junctions |
R1607 |
T5193 |
T5194 |
det |
the,sequences |
R1608 |
T5194 |
T5188 |
conj |
sequences,junctions |
R1609 |
T5195 |
T5194 |
prep |
at,sequences |
R1610 |
T5196 |
T5195 |
cc |
and,at |
R1611 |
T5197 |
T5195 |
conj |
around,at |
R1612 |
T5198 |
T5199 |
det |
the,sites |
R1613 |
T5199 |
T5197 |
pobj |
sites,around |
R1614 |
T5200 |
T5199 |
compound |
loxP,sites |
R1615 |
T5201 |
T5161 |
punct |
.,verified |