地被菊‘中国红’再生及遗传转化体系的建立

Establishment of Receptor and Genetic Transformation System of Ground-cover Chrysanthemum'China Red'

为建立地被菊‘中国红’的再生和遗传转化体系,以其无菌苗叶片为外植体,通过添加不同浓度的生长调节剂,研究其对诱导愈伤组织、不定芽的影响,并利用根瘤农杆菌介导法对地被菊‘中国红’叶片进行遗传转化,研究影响菊花转化的若干因素,建立一套高效的遗传转化体系。结果表明:地被菊‘中国红’在MS+2.0mg·L-16-BA+0.2mg·L-1NAA的培养基上获得了最高的不定芽分化率(89.6%);地被菊‘中国红’的最适生根培养基为1/2MS+0.3mg·L-1NAA,生根率达100%;其叶片外植体的遗传转化条件:OD600=0.6、农杆菌侵染时间为7min、共培养48h、延迟培养2d、卡那霉素筛选浓度为15mg·L-1、头孢霉素抑菌浓度为300mg·L-1。本研究为进一步开展菊花的基因工程育种奠定基础。

Different plant growth regulators that affect adventitious buds induction rate were studied in regeneration system of ground- cover chrysanthemum＇China Red＂ using the leaves of the aseptic seedings as explants. In order to establish an efficient genetic transformation system with leaves of ground-cover chrysanthemum＇China Red＇, several factors affected genetic transformation mediated by Agrobacterium were studied. The results showed that the highest rate of adventitious buds（89.6%） of ground-cover chrysanthemum＇China Red＂ was obtained on MS ＋ 2.0 mg·L^-1 6-BA ＋ 0.2 mg·L^-1NAA medium. The medium of 1/2MS＋0.3 mg·L^-1 NAA was the optimal for rooting, which had rooting rate of 100%. The conditions of genetic transformation： OD600=0.6, agrohacterium infection time for 7min, co-culture time for 48h, delayed screen time for 2 d, the kanamycin concentration was 15 mg·L^-1 and the cefsulodin concentration was 300 mg·L^-1. The establishment of high-efficiency transformation system laid a sound foundation for improving chrysanthemum by gene engineering.