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本帖最后由 zwb96 于 2013-5-9 23:41 编辑
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刚刚在Cell发表的文章（2013.5.9），转基因小鼠技术的鼻祖Rudolf Jaenisch领导的小组采用继ZFN,TALEN技术后最新的可修饰哺乳动物基因组的技术CRISPR以不可思议的高效率在小鼠胚胎干细胞一步敲除多个基因，更在短时间内通过合子内注射CRISPR系统（Cas9蛋白和sg-RNAs)建立了双基因敲除的小鼠模型，理论上只需4周时间就可完成，对基因敲除的技术来说是一次很大的飞跃，因为采用传统的同源重组的方法一般要9-15个月才能得到单个基因敲除的小鼠，敲除两个基因需要两年甚至更长的时间。以下是文章摘要。6 H. Z7 d( E2 O9 T+ w% S: I0 `
Mice carrying mutations in multiple genes are traditionally0 ^2 R# q# B5 S% F7 U% N
generated by sequential recombination in0 g- k' ~. _; S, e
embryonic stem cells and/or time-consuming intercrossing0 r2 g" r' @# c- S8 N
of mice with a single mutation. The
- E# \9 i8 M8 @, _7 ZCRISPR/Cas system has been adapted as an efficient
+ `" F1 a9 f7 o! ^ c+ Vgene-targeting technology with the potential
8 t% f( F* d5 d+ {' yfor multiplexed genome editing. We demonstrate
9 }4 k) o: J9 Ethat CRISPR/Cas-mediated gene editing allows the
' j( V; H: A }7 v( nsimultaneous disruption of five genes (Tet1, 2, 3,* N$ k0 ^, t4 g a
Sry, Uty - 8 alleles) in mouse embryonic stem (ES)
. u' W; X5 F$ G# ~/ H D& bcells with high efficiency. Coinjection of Cas9, T9 K8 ~- B" \$ z8 D$ Q+ H
mRNA and single-guide RNAs (sgRNAs) targeting& }8 N- @0 @" x1 R* C3 M3 R
Tet1 and Tet2 into zygotes generated mice with biallelic
9 w. t" S$ Y; g- bmutations in both genes with an efficiency of
8 K* ~% A3 f2 \- ]80%. Finally, we show that coinjection of Cas9. t( T8 ?7 l; F, v1 p
mRNA/sgRNAs with mutant oligos generated precise
2 X! b% f( X a3 a. z+ m2 [) B- Rpoint mutations simultaneously in two target8 s3 C8 _. g0 K) X3 L+ ]( ~* |' U
genes. Thus, the CRISPR/Cas system allows the: g5 S% o9 H- x4 ?8 b" | U
one-step generation of animals carrying mutations3 Z8 W" c( Q+ ` d
in multiple genes, an approach that will greatly accelerate' O" [2 K$ t) _9 f+ ~2 \
the in vivo study of functionally redundant
' N9 @5 i- Y* Z7 B. y5 |4 g9 zgenes and of epistatic gene interactions. |
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发表于 2013-5-9 23:38
发表于 2013-5-10 10:26
