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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。
: x, t: M3 j D6 y& rMice carrying mutations in multiple genes are traditionally0 f5 V# d+ }! _5 g
generated by sequential recombination in8 g( Z9 t6 t2 A
embryonic stem cells and/or time-consuming intercrossing
! l! P$ Q4 z- R2 eof mice with a single mutation. The
F& A$ ~4 ~6 c# D* zCRISPR/Cas system has been adapted as an efficient
) U: t1 s0 p2 x/ I& jgene-targeting technology with the potential
5 I$ {* Y9 e8 _" m7 V$ X$ a' \9 n E) S. {for multiplexed genome editing. We demonstrate
) `8 r8 V& r. H' H; y p, h2 qthat CRISPR/Cas-mediated gene editing allows the& @. R1 R4 n8 _( a% }3 \- X- C. d
simultaneous disruption of five genes (Tet1, 2, 3,' t7 W9 s$ g) P7 F4 d
Sry, Uty - 8 alleles) in mouse embryonic stem (ES)
8 c0 u7 O9 l4 e. {2 W" c7 Rcells with high efficiency. Coinjection of Cas9
" g' q0 x" h9 AmRNA and single-guide RNAs (sgRNAs) targeting
' U6 ?3 {, z8 [' q* d& {Tet1 and Tet2 into zygotes generated mice with biallelic; a- P1 o6 k/ u6 e1 x/ W, n* h
mutations in both genes with an efficiency of
; l, S1 }. u2 t) m80%. Finally, we show that coinjection of Cas9
% V9 c4 c# n4 v' ]1 FmRNA/sgRNAs with mutant oligos generated precise
# n* I0 e& S `- q8 ]# }point mutations simultaneously in two target
* i! W/ ]! I+ v7 ?" ]) T+ Hgenes. Thus, the CRISPR/Cas system allows the s% J! Y: o1 b4 ~, \9 q+ d
one-step generation of animals carrying mutations" X! ?6 p0 I8 u7 {" k T% B3 u
in multiple genes, an approach that will greatly accelerate
5 v" ^0 B' K2 ?0 vthe in vivo study of functionally redundant
/ J( n' U' e$ F! A/ Q2 t+ Jgenes 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
