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本帖最后由 zwb96 于 2013-5-9 23:41 编辑
# a3 @3 g2 Q( Z8 [% w m7 z' a- D0 ]8 t4 C- Z# z! D- p
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。
7 V2 I3 U. L# P; ^8 H) F, `Mice carrying mutations in multiple genes are traditionally$ i/ D# d' X2 @
generated by sequential recombination in
1 `; x2 x2 o$ b) Membryonic stem cells and/or time-consuming intercrossing
, [# E3 U: r* O1 {' N2 D; _of mice with a single mutation. The; D3 d+ j" w7 d4 H9 }
CRISPR/Cas system has been adapted as an efficient
+ U J3 c3 B7 ngene-targeting technology with the potential
. s; _( r* _" D8 s% e, f, d7 a9 R, Gfor multiplexed genome editing. We demonstrate
1 k2 s7 o9 k q$ C% G# i, m2 lthat CRISPR/Cas-mediated gene editing allows the
/ I- c7 q' V! x' bsimultaneous disruption of five genes (Tet1, 2, 3,) `" p+ f: Q) `$ p0 I* u8 `! {
Sry, Uty - 8 alleles) in mouse embryonic stem (ES)$ z7 h/ m0 g- G4 e
cells with high efficiency. Coinjection of Cas9- f/ B5 r! m3 ^
mRNA and single-guide RNAs (sgRNAs) targeting
! k, @4 K* }5 t- |/ xTet1 and Tet2 into zygotes generated mice with biallelic& l. J! p8 P+ }7 n9 |$ B
mutations in both genes with an efficiency of$ E0 @5 |; ~% I- u
80%. Finally, we show that coinjection of Cas9
6 T6 F. Q, M! C4 C' A8 umRNA/sgRNAs with mutant oligos generated precise! D5 a4 J$ {1 m& x/ B. }$ Z
point mutations simultaneously in two target
5 n: n* b8 Y' @- ]+ i) b/ Ngenes. Thus, the CRISPR/Cas system allows the
) x1 R8 o) o. _0 f( r! Hone-step generation of animals carrying mutations
6 ]+ P: P3 q/ f6 A- C/ {* M' V+ ain multiple genes, an approach that will greatly accelerate
! T+ q, `' i5 L# Fthe in vivo study of functionally redundant9 i) |* c2 K" j$ ~( I/ h
genes 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
