华南生态型黄瓜‘CU2’遗传转化体系的优化

Optimization of the Genetic Transformation System for the Ecotype Cucumber‘CU2’in South China

黄瓜是一种具有重要经济和生物价值的蔬菜作物。基因组编辑技术的广泛应用推动了黄瓜育种改良和基因组功能表征研究的快速发展。目前华南生态型黄瓜品种仍缺乏具有普遍适用性农杆菌介导的遗传转化体系,这限制了基因组编辑技术的高效应用。本研究以华南生态型黄瓜品种‘CU2’为材料,CsMATL基因编辑转基因株创制为例,进行遗传转化体系各种因素的优化,实现了该黄瓜品种外植体再分化芽的最高遗传转化率。研究结果表明选用SGM培养基,添加2 mg/L 6-BA、1 mg/L ABA后进行播种,在200μmol/L AS浓度下侵染外植体,共培养后采用摇床以及两次200 mg/L TMT溶液和三次无菌水清洗脱菌,之后在含有100 mg/L Kan、2 mg/L 6-BA、1 mg/L ABA的分化培养基中进行分化培养,利用上述优化方案可将‘CU2’的遗传转化率提高至3.19%。该研究为华南生态型黄瓜遗传转化体系的优化和功能基因操作奠定了理论基础。

Cucumber is a vegetable crop with significant economic and biological values.The widespread application of genome editing technology has promoted the rapid development of cucumber breeding and improvement,as well as researches on functional genomics.At present,the South China ecotype cucumber germplasm still lack a universally applicable Agrobacterium-mediated genetic transformation system,which limits the efficient application of genome editing technology.This study used the South China ecotype cucumber germplasm'CU2'as the material and the creation of CsMATL gene-edited transgenic plants as an example to optimize several factors in the genetic transformation system,achieving the highest genetic transformation rate of explant differentiation buds.The results showed that the operations of sowing the seeds on the SGM medium with 2 mg/L 6-BA and 1 mg/L ABA,co-culturing the explants at a concentration of 200μmol/L AS,and removing bacteria using two 200 mg/L TMT washes and three sterile water washes on a shaker yielded the positive results.Then,the differentiation culture was carried out in a differentiation medium containing 100 mg/L Kan,2 mg/L 6-BA,and 1 mg/L ABA.The genetic transformation rate of'CU2'can be increased to 3.19%by using the optimized method.This study laid a theoretical foundation for optimizing genetic transformation system and manipulating functional genes in South China ecotype cucumber.