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本帖最后由 zwb96 于 2013-5-9 23:41 编辑 + v4 V" q. J, Q/ g
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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。/ i5 I! W; D3 c# [) t/ {% t: v
Mice carrying mutations in multiple genes are traditionally
. E( p* R- s9 y0 igenerated by sequential recombination in" |5 e2 H/ w$ |' B- `( E1 e
embryonic stem cells and/or time-consuming intercrossing7 H0 d/ S( x2 ?4 O& ]
of mice with a single mutation. The
: [( ~' j7 S5 Y/ ICRISPR/Cas system has been adapted as an efficient; @; W6 M# M( E! ~7 {& d
gene-targeting technology with the potential5 l- a, ~* W# _8 Z: x
for multiplexed genome editing. We demonstrate
2 K+ x& Y6 {) p+ p. j% Sthat CRISPR/Cas-mediated gene editing allows the
n( z. m3 s( N. C$ Wsimultaneous disruption of five genes (Tet1, 2, 3,
. v& ]( S& e3 B- q; lSry, Uty - 8 alleles) in mouse embryonic stem (ES)
' A+ Z; D9 L( g1 R" m$ bcells with high efficiency. Coinjection of Cas9
y7 X6 Z/ @6 q( N0 f% P+ mmRNA and single-guide RNAs (sgRNAs) targeting. x4 [. B; p& `" s% j/ ~
Tet1 and Tet2 into zygotes generated mice with biallelic. d+ ]* R8 W& `1 Y8 ?, d, U( h
mutations in both genes with an efficiency of
9 G! Y' R5 w- f! q0 e80%. Finally, we show that coinjection of Cas9
; ?) A1 \7 r6 X* ~+ c. k7 M- g4 X& bmRNA/sgRNAs with mutant oligos generated precise
) u: f9 A% O; ipoint mutations simultaneously in two target
# h4 `% R V( b5 fgenes. Thus, the CRISPR/Cas system allows the* i+ H+ _5 ]1 _& V% n
one-step generation of animals carrying mutations M+ ~, Y3 L- L8 ]& ~
in multiple genes, an approach that will greatly accelerate( T; T8 [/ Q( b* d
the in vivo study of functionally redundant
) _: T/ l* {4 |- t, ^3 `9 [genes and of epistatic gene interactions. |
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发表于 2013-5-9 23:38
发表于 2013-7-19 10:47
