基于CRISPR RNA/Cas9系统的基因组定点修饰技术.

Cas9 是一种核酸内切酶,CRISPR RNA用来引导这个酶去切割目标DNA.目标怎么设定?既然这里有RNA,那么

它可以去发现和其互补的DNA.所谓定点靠这个来实现.

这个技术的妙出在于:http://yunpan.cn/QGrT5k5BFXnbk

1 获得的最终产品是经过了遗传修饰的(定点修饰,通过转基因的方法定点修饰,因为这个系

统是细菌里的,所以要靠转基因的方法转到植物中),但终产品里却可以不包含任何转基因的成分(CRISPR

RNA/Cas9,构建到一个载体里,转基因植株以杂合的形式保存就行了).

2 这个技术修饰的是一个物种自身的基因,所以没有明显可见的安全性问题.

3 呵呵,这个技术和我前面设想的用水稻的基因来改良水稻的观点是吻合的.(http://blog.sciencenet.cn/blog-479743-709708.html)

4 外行人,不知道基因还可以这样转.

新东西我也不是很熟,下面是在Pubmed搜索的一些文献,有兴趣的朋友可以细致的看看.估计明年的国家自然科

学基金会有人写这个.

我下载到的一些全文愿意分享:
1: Walsh RM, Hochedlinger K. A variant CRISPR-Cas9 system adds versatility to
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2: Chen C, Fenk LA, de Bono M. Efficient genome editing in Caenorhabditis elegans
by CRISPR-targeted homologous recombination. Nucleic Acids Res. 2013 Sep 5. [Epub
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3: Kondo S, Ueda R. Highly Improved Gene Targeting by Germline-Specific Cas9
Expression in Drosophila. Genetics. 2013 Sep 3. [Epub ahead of print] PubMed
PMID: 24002648.

4: Jiang W, Zhou H, Bi H, Fromm M, Yang B, Weeks DP. Demonstration of
CRISPR/Cas9/sgRNA-mediated targeted gene modification in Arabidopsis, tobacco,
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5: Yu Z, Ren M, Wang Z, Zhang B, Rong YS, Jiao R, Gao G. Highly Efficient Genome
Modifications Mediated by CRISPR/Cas9 in Drosophila. Genetics. 2013
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6: Pattanayak V, Lin S, Guilinger JP, Ma E, Doudna JA, Liu DR. High-throughput
profiling of off-target DNA cleavage reveals RNA-programmed Cas9 nuclease
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2013 Aug 11. PubMed PMID: 23934178.

7: Mali P, Aach J, Stranges PB, Esvelt KM, Moosburner M, Kosuri S, Yang L, Church
GM. CAS9 transcriptional activators for target specificity screening and paired
nickases for cooperative genome engineering. Nat Biotechnol. 2013
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8: Hsu PD, Scott DA, Weinstein JA, Ran FA, Konermann S, Agarwala V, Li Y, Fine
EJ, Wu X, Shalem O, Cradick TJ, Marraffini LA, Bao G, Zhang F. DNA targeting
specificity of RNA-guided Cas9 nucleases. Nat Biotechnol. 2013 Sep;31(9):827-32.
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9: Dickinson DJ, Ward JD, Reiner DJ, Goldstein B. Engineering the Caenorhabditis
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10: Fujii W, Kawasaki K, Sugiura K, Naito K. Efficient generation of large-scale
genome-modified mice using gRNA and CAS9 endonuclease. Nucleic Acids Res. 2013
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11: Ran FA, Hsu PD, Lin CY, Gootenberg JS, Konermann S, Trevino AE, Scott DA,
Inoue A, Matoba S, Zhang Y, Zhang F. Double Nicking by RNA-Guided CRISPR Cas9 for
Enhanced Genome Editing Specificity. Cell. 2013 Aug 28. doi:pii:
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12: Yang H, Wang H, Shivalila CS, Cheng AW, Shi L, Jaenisch R. One-Step
Generation of Mice Carrying Reporter and Conditional Alleles by
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13: Cheng AW, Wang H, Yang H, Shi L, Katz Y, Theunissen TW, Rangarajan S,
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14: Waaijers S, Portegijs V, Kerver J, Lemmens BB, Tijsterman M, van den Heuvel
S, Boxem M. CRISPR/Cas9-Targeted Mutagenesis in Caenorhabditis elegans. Genetics.
2013 Aug 26. [Epub ahead of print] PubMed PMID: 23979586.

15: Tzur YB, Friedland AE, Nadarajan S, Church GM, Calarco JA, Colaiácovo MP.
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CRISPR-Cas9 System. Genetics. 2013 Aug 26. [Epub ahead of print] PubMed PMID:
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16: Katic I, Großhans H. Targeted Heritable Mutation and Gene Conversion by
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17: Chiu H, Schwartz HT, Antoshechkin I, Sternberg PW. Transgene-Free Genome
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18: Cho SW, Lee J, Carroll D, Kim JS, Lee J. Heritable Gene Knockout in
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19: Jao LE, Wente SR, Chen W. Efficient multiplex biallelic zebrafish genome
editing using a CRISPR nuclease system. Proc Natl Acad Sci U S A. 2013 Aug
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23918387; PubMed Central PMCID: PMC3752207.

20: Xie K, Yang Y. RNA-guided Genome Editing in Plants Using A CRISPR-Cas System.
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21: Lo TW, Pickle CS, Lin S, Ralston EJ, Gurling M, Schartner CM, Bian Q, Doudna
JA, Meyer BJ. Precise and Heritable Genome Editing in Evolutionarily Diverse
Nematodes Using TALENs and CRISPR/Cas9 to Engineer Insertions and Deletions.
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22: Hou Z, Zhang Y, Propson NE, Howden SE, Chu LF, Sontheimer EJ, Thomson JA.
Efficient genome engineering in human pluripotent stem cells using Cas9 from
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23: Fujita T, Fujii H. Efficient isolation of specific genomic regions and
identification of associated proteins by engineered DNA-binding molecule-mediated
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24: Cradick TJ, Fine EJ, Antico CJ, Bao G. CRISPR/Cas9 systems targeting β-globin
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25: Gratz SJ, Cummings AM, Nguyen JN, Hamm DC, Donohue LK, Harrison MM, Wildonger
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26: Friedland AE, Tzur YB, Esvelt KM, Colaiácovo MP, Church GM, Calarco JA.
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27: Bikard D, Jiang W, Samai P, Hochschild A, Zhang F, Marraffini LA.
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28: Xiao A, Wang Z, Hu Y, Wu Y, Luo Z, Yang Z, Zu Y, Li W, Huang P, Tong X, Zhu
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29: Yang L, Guell M, Byrne S, Yang JL, De Los Angeles A, Mali P, Aach J,
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30: Perez-Pinera P, Kocak DD, Vockley CM, Adler AF, Kabadi AM, Polstein LR,
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33: Wang H, Yang H, Shivalila CS, Dawlaty MM, Cheng AW, Zhang F, Jaenisch R.
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Plant genome editing made easy: targeted mutagenesis in model and crop plants using the CRISPR/Cas system