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利用CRISPR/Cas9造成的大片段缺失,先正达公司创造了高产籼稻

2017-06-10来源:莱肯生物
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      利用CRISPR/Cas9技术进行基因组编辑成为作物育种的一个有力手段。大多数情况下,CRISPR/Cas9基因编辑导致的基因组编辑形式是产生碱基替换或小片段的插入缺失(InDels),大片段缺失的情况极少见。最近,先正达生物科技(中国)有限公司的研发人员利用CRISPR/Cas9技术造成了籼稻中DEP1基因的大片段缺失。

      DEP1基因(DENSE AND ERECT PANICLE1)是中科院遗传发育所傅向东课题组克隆的产量性状基因。粳稻中存在着显性dep1突变基因,能促进细胞分裂,使得植株半矮化、稻穗变密、枝梗数增加和每穗籽粒数增多,从而促使水稻增产。然而,籼稻中并不存在dep1突变基因。如果利用常规育种方法将粳稻中的dep1转育到籼稻中将十分耗时耗力。因此,先正达的研发人员尝试利用CRISPR/Cas9技术对籼稻中的DEP1基因进行突变。他们成功造成了DEP1区域基因组上10kb片段的缺失,而且编辑效率超过9%,并得到了具有增产潜力的编辑后水稻材料。
      该研究不仅对高产水稻培育工作提供了更高效的方法,高效的大片段缺失编辑也将扩展基因编辑技术的应用范围。
      先正达生物科技(中国)有限公司是先正达全球七大研究中心之一,2008年落户北京中关村生命科学园,同年10月正式运营,总投资超过1亿美元。现有约100人的研发团队,专注于玉米、大豆、水稻等主要作物早期生物技术和天然农艺性状领域的研究,提高作物产量、抗旱性及抗病抗虫能力。
Plant Cell Rep. 2017 Jun 5.
Deletion of a target gene in Indica rice via CRISPR/Cas9.
Author
Wang Y, Geng L, Yuan M……Chai Z, Fu X, Li X*.
*: Syngenta Biotechnology China.
Abstract
CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 is a rapidly developing technology used to produce gene-specific modifications in both mammalian and plant systems. Most CRISPR-induced modifications in plants reported to date have been small insertions or deletions. Few large target gene deletions have thus far been reported, especially for Indica rice. In this study, we designed multiple CRISPR sgRNAs and successfully deleted DNA fragments in the gene DENSE AND ERECT PANICLE1 (DEP1) in the elite Indica rice line IR58025B. We achieved deletion frequencies of up to 21% for a 430 bp target and 9% for a 10 kb target among T0 events. Constructs with four sgRNAs did not generate higher full-length deletion frequencies than constructs with two sgRNAs. The multiple mutagenesis frequency reached 93% for four targets, and the homozygous mutation frequency reached 21% at the T0 stage. Important yield-related trait characteristics, such as dense and erect panicles and reduced plant height, were observed in dep1 homozygous T0 mutant plants produced by CRISPR/Cas9. Therefore, we successfully obtained deletions in DEP1 in the Indica background using the CRISPR/Cas9 editing tool at relatively high frequency.

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