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本帖最后由 zwb96 于 2013-5-9 23:41 编辑
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/ }: W: C) j5 C3 q" h% F. v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。2 F# U/ a( S7 o$ |$ v! G& k) t5 q
Mice carrying mutations in multiple genes are traditionally
/ O, m9 F" H: W0 _. x$ k8 Rgenerated by sequential recombination in2 U0 m! s% L" r5 e
embryonic stem cells and/or time-consuming intercrossing [$ }# V) Y- C* Y, n. G1 Q
of mice with a single mutation. The' r$ y1 W. x* B2 N# P
CRISPR/Cas system has been adapted as an efficient) j$ j) ~: V7 {& u: v
gene-targeting technology with the potential! M0 t) w/ [8 u# C4 I
for multiplexed genome editing. We demonstrate
' _2 o" y; b9 _7 z- l, p# Ithat CRISPR/Cas-mediated gene editing allows the6 u) A0 ~$ K* w( {0 l- j
simultaneous disruption of five genes (Tet1, 2, 3,
- Z' M# b& A3 `5 Q# n3 H3 n5 T, ESry, Uty - 8 alleles) in mouse embryonic stem (ES)7 [$ |+ p' ^' Y1 B F
cells with high efficiency. Coinjection of Cas9
P0 u2 @* D( X; G# S! i- J( mmRNA and single-guide RNAs (sgRNAs) targeting
# y2 b. N2 w7 [( qTet1 and Tet2 into zygotes generated mice with biallelic7 |) l# u# j: V. O
mutations in both genes with an efficiency of/ o D. r2 d, j2 J* `$ H& {
80%. Finally, we show that coinjection of Cas90 P, z3 x) d' s. R( u" Z, ?
mRNA/sgRNAs with mutant oligos generated precise. S7 H9 _8 y! {
point mutations simultaneously in two target- }# ~% K9 {/ I4 S: `7 a- r! ~4 ^% P
genes. Thus, the CRISPR/Cas system allows the3 z6 b( l8 K0 Y _& w( \5 \
one-step generation of animals carrying mutations0 k% x( ?9 L4 r7 q
in multiple genes, an approach that will greatly accelerate% w4 @* l. E1 H! @8 `
the in vivo study of functionally redundant" O1 }8 S. `* r
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
