Fu Yongyao, Shu Liling, Li Hanyi, Zhang Xingming, Liu Xuan, Ou Zhengying, Liang Xiaomeng, Qi Xiangying, Yang Liping
School of Advanced Agriculture and Bioengineering, Yangtze Normal University, Chongqing 408100, China.
School of Life Sciences, Yan'an University, Yan'an 716000, China.
Plants (Basel). 2023 May 16;12(10):1992. doi: 10.3390/plants12101992.
var. , commonly called Longya lily, is a well-known flower and vegetable plant in China that has poor tolerance to fungal disease. The molecularimprovement has mainly been restricted to an efficient regeneration and transformation system. In this study, the highly efficient regeneration of Longya lily was established through the optimization of embryogenic callus, adventitious shoot and rooting induction. The major factors influencing transformation (antibiotics, concentration, infection time, suspension solution and coculture medium) were examined. The expression responses of PR promoters ( and ) to were assessed in transgenic calli. The results showed that Murashige and Skoog (MS) medium with 1.0 mg·L picloram (PIC) and 0.2 mg·L 1-naphthaleneacetic acid (NAA) under light conditions and MS with 0.5 mg·L 6-benzylaminopurine (6-BA) and 1.0 mg·L NAA under darkness were optimal for embryogenic callus induction (64.67% rate) and proliferation (3.96 coefficient). Callus inoculation into MS containing 2.0 mg·L thidiazuron (TDZ), 0.4 mg·L NAA, 1.0 mg·L TDZ and 0.5 mg·L NAA led to shooting induction (92.22 of rate) and proliferation (3.28 of coefficient) promotion, respectively. The rooting rate reached 99.00% on MS with 0.3 mg·L NAA. Moreover, a transformation rate of 65.56% was achieved by soaking the callus in at an OD of 0.4 for 10 min in modified MS without NHNO as the suspension solution and coculture medium before selecting 75 mg·L hygromycin and 300 mg·L cefotaxime. Only the promoter was obviously expressed in transgenic calli. These results could facilitate the generation of Longya lily transgenic plants with improved resistance.
龙牙百合变种，通常被称为龙牙百合，是中国一种著名的花卉和蔬菜作物，但对真菌病害的耐受性较差。分子改良主要局限于高效的再生和转化系统。在本研究中，通过优化胚性愈伤组织、不定芽和生根诱导，建立了龙牙百合的高效再生体系。研究了影响转化的主要因素（抗生素、浓度、感染时间、悬浮液和共培养基）。在转基因愈伤组织中评估了病程相关蛋白（PR）启动子（和）对的表达响应。结果表明，在光照条件下，添加1.0 mg·L 氯吡脲（PIC）和0.2 mg·L 1-萘乙酸（NAA）的Murashige和Skoog（MS）培养基，以及在黑暗条件下添加0.5 mg·L 6-苄基腺嘌呤（6-BA）和1.0 mg·L NAA的MS培养基，最适合胚性愈伤组织诱导（诱导率64.67%）和增殖（增殖系数3.96）。将愈伤组织接种到含有2.0 mg·L噻苯隆（TDZ）、0.4 mg·L NAA、1.0 mg·L TDZ和0.5 mg·L NAA的MS培养基中，分别促进了不定芽诱导（诱导率92.22%）和增殖（增殖系数3.28）。在添加0.3 mg·L NAA的MS培养基上生根率达到99.00%。此外，在不添加NHNO的改良MS培养基中，将愈伤组织在OD为0.4的菌液中浸泡10 min作为悬浮液和共培养基，然后选择75 mg·L潮霉素和300 mg·L头孢噻肟，转化效率达到65.56%。在转基因愈伤组织中只有启动子明显表达。这些结果有助于培育出具有改良抗性的龙牙百合转基因植株。
