基于CRISPR/Cas9技术创建甜菜BvCENH3基因突变体的研究

doi:10.12190/j.issn.2096-1197.2023.02.01

Abstract

Abstract: 【Objective】The CRISPR/Cas9 technology was used to edit the BvCENH3 gene in sugar beet aiming to establish an efficient genome editing system.【Methods】Taking sugar beet BvCENH3 gene as the editing target,double candidate targets were selected to construct gene editing vector. Transgenic sugar beet plants were produced through agrobacterium-mediated transformation. Next generation sequencing technology was utilized to identify the mutation types and droplet digital PCR was employed to screen low-copy mutant plants.【Results】82 transgenic sugar beet plants were obtained,40 of which were successfully edited,with an editing efficiency of 48.78%. Target 1 was more efficient than target 2. There were five mutation types,including single base substitution（T→G、A→C） and base deletion（TC、TCTC deletion）. 23 low-copy edited plants were selected,with BvCENH3 insertion copy numbers ranging from 1.1 to 1.9.【Conclusion】Successfully edited BvCENH3 in sugar beet with 40 BvCENH3 gene mutants obtained. Preliminary established sugar beet genome editing system,laying theoretical and technical foundations for sugar beet haploid breeding.

Key words: Sugar beet, BvCENH3, CRISPR/Cas9, Genome editing, Mutants

CLC Number:

S566.335.3

HAN Ping′an, TANG Kuan′gang, CHANG Yue, SUN Ruifen, WANG Liang, ZHANG Ziqiang, FU Zengjuan, ZHAO Shangmin, WU Xinrong, LI Xiaodong. Research on the construction of sugar beet BvCENH3 gene mutants based on CRISPR/Cas9 technology[J].Journal of Northern Agriculture, 2023, 51(2): 1-11.

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Research on the construction of sugar beet BvCENH3 gene mutants based on CRISPR/Cas9 technology

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