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Cell:张锋发表综述详细介绍第二类CRISPR-Cas系统

2017-02-16 MedSci MedSci原创

近期张锋教授也与另外两位学者在Cell杂志上发表了题为“SnapShot: Class 2 CRISPR-Cas Systems”的特写文章,介绍了新一代CRISPR基因组编辑系统:Class 2 CRISPR-Cas Systems。 2015年,张锋及其同事们就报告称发现了一种不同的CRISPR系统,具有潜力实现更简单、更精确的基因组工程操作。这个新系统是通过在不同类型的细菌中搜寻了成百上千

近期张锋教授也与另外两位学者在Cell杂志上发表了题为“SnapShot: Class 2 CRISPR-Cas Systems”的特写文章,介绍了新一代CRISPR基因组编辑系统:Class 2 CRISPR-Cas Systems。

2015年,张锋及其同事们就报告称发现了一种不同的CRISPR系统,具有潜力实现更简单、更精确的基因组工程操作。这个新系统是通过在不同类型的细菌中搜寻了成百上千种的CRISPR系统,寻找具有有用特性的酶,结果来自氨基酸球菌属(Acidaminococcus)和毛螺菌科(Lachnospiraceae)的Cpf1酶成为新的候选物。

这一新发现的Cpf1系统有几个重要的方面不同于以往描述的Cas9,在生物通最先报道的张锋Cell:新一代CRISPR基因组编辑系统这篇文章中就提到:Cpf1系统更简单一些,它只需要一条RNA。Cpf1酶也比标准SpCas9要小,使得它更易于传送至细胞和组织内;Cpf1以一种不同于Cas9的方式切割DNA。当Cas9复合物切割DNA时,它切割的是同一位点的两条链,留下的“平端”(blunt ends)在重新连接时往往会发生突变。采用Cpf1复合物生成的两条链切口是偏移的,在裸露端留下了短悬端(overhang)。这预计有助于精确插入,使得研究人员能够更有效及精确地整合一段DNA;Cpf1切口远离识别位点,这意味着即便在切割位点靶基因突变,仍然可以进行再度切割,提供了多次机会来校正编辑;Cpf1系统为选择靶位点提供了新的灵活性。像Cas9一样,Cpf1复合物必须首选附着PAM,短序列,选择的靶点靠近自然存在的PAM序列。Cpf1系统识别的PAM序列与Cas9截然不同。这在靶向某些基因组如疟原虫及人类基因组时可能是个优势。

这篇最新综述则更加概括的描述了这一系统,见下图:

文章指出,第二类CRISPR-Cas系统基于不同的效应蛋白家族,可以分为3种类型和9种亚型,其中譬如Cas9和Cas12a(Cpf1)已成功地用于基因组工程。在此前的研究中,张锋研究组也采用了一种新生物信息学方法来发现暂时被命名为C2c1、C2c2和C2c3的新蛋白,他们开发出一系列的计算方法来搜索NIH基因组数据库,鉴别新的CRISPR-Cas系统(详细见:Cell:第二代基因编辑神器诞生,CRISPR/Cpf1让一切皆有可能!)。

除此之外,张锋研究组也提出了一些新的尝试,如他们构建出了32个具有单个点突变的Cas9突变体,借助于已验证易错配的向导RNA将它们靶向EMX1基因。张锋将这些特异性增强型Cas9蛋白称作为“eSpCas9变体”。测试大量的向导RNAs与具单个点突变的eSpCas9和具三个点突变的eSpCas9,他们发现两种eSpCas9能够以相似的效率编辑靶位点。

张锋课题组还开发了一些基于功能获得性CRISPR的筛查方法。利用这一基于CRISPR的新工具来激活SAM基因,他的研究小组证实这一工具可以激活采用旧方法难于开启的12种不同的基因。“有许多基因采用旧系统无法激活,这一新系统能够激活转录达100倍或1000倍,”张锋说。

去年年底,张锋研究组发现Cpf1能够加工自己的CRISPR RNA (crRNA)。而且Cpf1介导的pre-crRNA加工是独立于DNA剪切的。这种能力可以用来简化多重基因组编辑。他们在此基础上设计CRISPR阵列,用一个载体同时在哺乳动物细胞编辑了四个基因,在小鼠大脑编辑了三个基因。这是在CRISPR–Cpf1的基础上打造了一个多重化基因编辑系统。

原始出处:

Makarova KS, Zhang F, Koonin EV. SnapShot: Class 2 CRISPR-Cas Systems. Cell. 2017 Jan 12;168(1-2):328-328.e1. doi: 10.1016/j.cell.2016.12.038.

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    2017-03-27 维他命
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    2017-02-18 yuandd
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    2017-02-18 yaanren
  6. [GetPortalCommentsPageByObjectIdResponse(id=1960027, encodeId=6e8f196002ed0, content=<a href='/topic/show?id=dbe6445416' target=_blank style='color:#2F92EE;'>#CEL#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=80, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=4454, encryptionId=dbe6445416, topicName=CEL)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=c3ff68, createdName=维他命, createdTime=Mon Mar 27 04:40:00 CST 2017, time=2017-03-27, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1869768, encodeId=12651869e68f8, content=<a href='/topic/show?id=fd764459a8' target=_blank style='color:#2F92EE;'>#Cell#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=65, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=4459, encryptionId=fd764459a8, topicName=Cell)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=524d95, createdName=zhaozhouchifen, createdTime=Tue Jun 13 01:40:00 CST 2017, time=2017-06-13, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1275994, encodeId=cf0112e59944e, content=<a href='/topic/show?id=755d5211a0' target=_blank style='color:#2F92EE;'>#CRISPR#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=64, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=5211, encryptionId=755d5211a0, topicName=CRISPR)], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=583c133, createdName=yuandd, createdTime=Sat Feb 18 12:40:00 CST 2017, time=2017-02-18, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1455273, encodeId=7fcf14552e3f3, content=<a href='/topic/show?id=ce8452137f' target=_blank style='color:#2F92EE;'>#CRISPR-Cas#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=71, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=5213, encryptionId=ce8452137f, topicName=CRISPR-Cas)], attachment=null, authenticateStatus=null, createdAvatar=null, createdBy=e8505649833, createdName=guihongzh, createdTime=Sat Feb 18 12:40:00 CST 2017, time=2017-02-18, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=1624816, encodeId=ca7c162481687, content=<a href='/topic/show?id=0333412062' target=_blank style='color:#2F92EE;'>#Cas#</a>, beContent=null, objectType=article, channel=null, level=null, likeNumber=54, replyNumber=0, topicName=null, topicId=null, topicList=[TopicDto(id=4120, encryptionId=0333412062, topicName=Cas)], attachment=null, authenticateStatus=null, createdAvatar=null, createdBy=411420689333, createdName=yaanren, createdTime=Sat Feb 18 12:40:00 CST 2017, time=2017-02-18, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=176596, encodeId=b4d31e659602, content=好好努力, beContent=null, objectType=article, channel=null, level=null, likeNumber=118, replyNumber=0, topicName=null, topicId=null, topicList=[], attachment=null, authenticateStatus=null, createdAvatar=, createdBy=45101626056, createdName=忠诚向上, createdTime=Fri Feb 17 10:18:34 CST 2017, time=2017-02-17, status=1, ipAttribution=), GetPortalCommentsPageByObjectIdResponse(id=176536, encodeId=ea1a1e6536d4, content=阅读了谢谢分享, beContent=null, objectType=article, channel=null, level=null, likeNumber=112, replyNumber=0, topicName=null, topicId=null, topicList=[], attachment=null, authenticateStatus=null, createdAvatar=http://cdnapi.center.medsci.cn/medsci/head/2017/04/21/72142930668371951a41ef4ea4b233e6.jpg, createdBy=dcbc1715228, createdName=刘煜, createdTime=Fri Feb 17 06:18:53 CST 2017, time=2017-02-17, status=1, ipAttribution=)]
    2017-02-17 忠诚向上

    好好努力

    0

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    2017-02-17 刘煜

    阅读了谢谢分享

    0

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