High efficient multisites genome editing in allotetraploid cotton (Gossypium hirsutum) using CRISPR/Cas9 system

生物 基因组编辑 清脆的 Cas9 遗传学 基因组 桑格测序 基因 DNA测序 计算生物学
作者
Pengcheng Wang,Jun Zhang,Lin Sun,Yizan Ma,Jiao Xu,Sijia Liang,Jinwu Deng,Jiafu Tan,Zhang Qing-hua,Lili Tu,Henry Daniell,Shuangxia Jin,Xianlong Zhang
出处
期刊:Plant Biotechnology Journal [Wiley]
卷期号:16 (1): 137-150 被引量:217
标识
DOI:10.1111/pbi.12755
摘要

Gossypium hirsutum is an allotetraploid with a complex genome. Most genes have multiple copies that belong to At and Dt subgenomes. Sequence similarity is also very high between gene homologues. To efficiently achieve site/gene-specific mutation is quite needed. Due to its high efficiency and robustness, the CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 system has exerted broad site-specific genome editing from prokaryotes to eukaryotes. In this study, we utilized a CRISPR/Cas9 system to generate two sgRNAs in a single vector to conduct multiple sites genome editing in allotetraploid cotton. An exogenously transformed gene Discosoma red fluorescent protein2(DsRed2) and an endogenous gene GhCLA1 were chosen as targets. The DsRed2-edited plants in T0 generation reverted its traits to wild type, with vanished red fluorescence the whole plants. Besides, the mutated phenotype and genotype were inherited to their T1 progenies. For the endogenous gene GhCLA1, 75% of regenerated plants exhibited albino phenotype with obvious nucleotides and DNA fragments deletion. The efficiency of gene editing at each target site is 66.7-100%. The mutation genotype was checked for both genes with Sanger sequencing. Barcode-based high-throughput sequencing, which could be highly efficient for genotyping to a population of mutants, was conducted in GhCLA1-edited T0 plants and it matched well with Sanger sequencing results. No off-target editing was detected at the potential off-target sites. These results prove that the CRISPR/Cas9 system is highly efficient and reliable for allotetraploid cotton genome editing.
最长约 10秒,即可获得该文献文件

科研通智能强力驱动
Strongly Powered by AbleSci AI
更新
大幅提高文件上传限制,最高150M (2024-4-1)

科研通是完全免费的文献互助平台,具备全网最快的应助速度,最高的求助完成率。 对每一个文献求助,科研通都将尽心尽力,给求助人一个满意的交代。
实时播报
竹子发布了新的文献求助10
1秒前
小小完成签到,获得积分10
1秒前
上官若男应助hank采纳,获得10
3秒前
搜集达人应助廿一采纳,获得10
4秒前
学术狗发布了新的文献求助10
4秒前
6秒前
七七完成签到,获得积分10
7秒前
7秒前
Ron完成签到,获得积分10
7秒前
8秒前
看看发布了新的文献求助10
9秒前
俊逸的汲完成签到,获得积分10
11秒前
Fei发布了新的文献求助10
12秒前
学术狗完成签到,获得积分10
12秒前
14秒前
长发飘飘完成签到 ,获得积分10
14秒前
15秒前
17秒前
俊逸的汲发布了新的文献求助10
18秒前
19秒前
廿一完成签到,获得积分10
19秒前
najibveto应助lilililili采纳,获得10
20秒前
老詹头应助流年羽采纳,获得10
22秒前
dadadada发布了新的文献求助30
23秒前
24秒前
七七完成签到 ,获得积分10
24秒前
26秒前
昆仑山吴某完成签到 ,获得积分10
26秒前
27秒前
大个应助洛源采纳,获得10
32秒前
hfcao发布了新的文献求助10
33秒前
33秒前
LHH完成签到 ,获得积分10
34秒前
34秒前
JQK完成签到 ,获得积分10
34秒前
36秒前
quhayley应助dadadada采纳,获得10
37秒前
外向翠萱完成签到,获得积分20
38秒前
Neinei发布了新的文献求助10
38秒前
39秒前
高分求助中
Evolution 10000
Sustainability in Tides Chemistry 2800
юрские динозавры восточного забайкалья 800
English Wealden Fossils 700
An Introduction to Geographical and Urban Economics: A Spiky World Book by Charles van Marrewijk, Harry Garretsen, and Steven Brakman 600
Diagnostic immunohistochemistry : theranostic and genomic applications 6th Edition 500
Mantiden: Faszinierende Lauerjäger Faszinierende Lauerjäger 400
热门求助领域 (近24小时)
化学 医学 生物 材料科学 工程类 有机化学 生物化学 物理 内科学 纳米技术 计算机科学 化学工程 复合材料 基因 遗传学 催化作用 物理化学 免疫学 量子力学 细胞生物学
热门帖子
关注 科研通微信公众号,转发送积分 3153361
求助须知:如何正确求助?哪些是违规求助? 2804608
关于积分的说明 7860306
捐赠科研通 2462547
什么是DOI,文献DOI怎么找? 1310806
科研通“疑难数据库(出版商)”最低求助积分说明 629396
版权声明 601794