四川大学学报（自然科学版）2025，Vol.62Issue(2)：259-270,12.DOI:10.19907/j.0490-6756.240302
雌雄青杨组织培养和PcXTH22基因过表达及敲除
Tissue regeneration of male and female Populus cathayana and overexpression and knockdown of PcXTH22 gene
摘要
Abstract
To achieve genetic improvement of Populus cathayana Rehd through genetic engineering meth-ods,male and female poplar were used as materials to establish a systematic tissue culture regeneration.The optimal mediums for the differentiation,proliferation and rooting of female P.cathayana were MS+30 g/L sucrose+6 g/L agar+0.1 mg/L TDZ+0.3 mg/L IBA,MS+30 g/L sucrose+7 g/L agar+0.1 mg/L 6-BA+0.5 mg/L IBA,WPM+20 g/L sucrose+7 g/L agar+0.1 mg/L IBA+0.2 mg/L NAA;those for male P.cathayana were:WPM+30 g/L sucrose+6 g/L agar+0.1 mg/L TDZ+0.3 mg/L NAA,MS+30 g/L sucrose+7 g/L agar+1.0 mg/L 6-BA+0.1 mg/L NAA,WPM+20 g/L sucrose+7 g/L agar+0.2 mg/L IBA+0.02 mg/L NAA.Subsequently,the leaves of female P.cathayana after 1 month of sub-culture were used as genetic transformation receptor material for overexpression and knockout of the PcXTH22 gene.After infecting with Agrobacterium solution for 10～15 minutes,the leaf discs were co-cultured with Agrobacterium for 48 h and then transferred into mediacontaining 10 mg/L hygromycin and 200 mg/L temetine for selective culture.The obtained resistant plants were detected by PCR,and the trans-formation rate was almost 100%.Quantitative PCR showed that the transcriptional expression level of PcXTH22 in overexpressed strains increased by 40-824 times.Among the 5 CRISPR-Cas9-transformed lines,1 line was found to have frameshift mutation on both alleles and the protein translation was terminated prematurely,and the editing efficiency was 20%.Therefore,the study provides an excellent example of ge-netic improvement using our endemic native tree species.关键词
青杨/雌雄植株/组织再生/遗传转化体系/PcXTH22/CRISPR-Cas9基因编辑Key words
Populus cathayana/Dioecious/Tissue regeneration/Genetic transformation/PcXTH22/CRISPR-Cas9 gene edition分类
农业科技引用本文复制引用
陈遥,李婧,赖诗宜,张雪驰,陈凡,张胜..雌雄青杨组织培养和PcXTH22基因过表达及敲除[J].四川大学学报（自然科学版）,2025,62(2):259-270,12.基金项目
国家自然科学基金(32271830) （32271830）
青藏高原寒旱区大规格乔木移植关键技术研究及应用项目(2846G202400001) （2846G202400001）
