红蓝光促进陆地棉愈伤组织诱导和增殖

Red and Blue Light Promotes Cotton Callus Induction and Proliferation

【目的】陆地棉(Gossypium hirsutum)的遗传转化技术存在转化周期长、转化效率低等问题,其中,愈伤组织增殖速度慢是导致棉花转化周期长的关键因素。探究陆地棉愈伤组织增殖的最适光照条件,为缩短棉花转化周期提供技术基础。【方法】以陆地棉受体品种WC的下胚轴为外植体诱导愈伤组织,设置红光、蓝光、红蓝光(1﹕1)和白光(CK)4种不同光照处理,研究不同光质下愈伤组织的增殖速度、形态特征,以及相关基因表达水平和相关酶活性的差异,解析光照对愈伤组织增殖的影响并获得最优光照条件。【结果】不同光质对诱导棉花外植体出愈和愈伤组织生长具有明显影响。在4种不同光照处理下,红蓝光相较其他光质具有更显著地促进外植体出愈和愈伤组织增殖的效果,表现为愈伤组织分化时间更短,7—15 d,即与其他处理存在明显表型差异,7 d愈伤组织鲜重红蓝光(0.39 g)>蓝光(0.34 g)>白光(0.24 g)>红光(0.23 g),15 d时,红蓝光(1.15 g)>蓝光(0.98 g)>白光(0.69 g)>红光(0.51 g),红蓝光照射下的愈伤组织鲜重是白光对照的1.65倍,比单一蓝光或红光照射下的愈伤组织鲜重增加16.5%或125.5%;与此相一致的是,红蓝光处理下,第二周的愈伤增殖率也最高,达14.67%,比红光处理(7.17%)增加1倍多。4种光质下,愈伤增殖及体胚再生相关基因的表达量,红蓝光处理均显示最高。此外,红蓝光处理下愈伤组织的过氧化氢酶(CAT)活性显著提高,活性氧(ROS)含量低于白光对照。【结论】不同光质条件下,棉花外植体的愈伤组织增殖速度不同,红蓝光(1﹕1)促进外植体出愈和愈伤增殖表现最优,其次为蓝光,再次为白光,单独的红光不利于棉花出愈和愈伤增殖。红蓝光(1﹕1)处理诱导光受体及其互作因子、愈伤增殖等相关基因的表达,增强愈伤组织内过氧化氢酶的活性,降低活性氧含量,促进细胞分裂,进而促进愈伤增殖。

【Objective】Upland cotton(Gossypium hirsutum)genetic transformation faces series of challenges such as a prolonged cycle and low efficiency,with the relatively slow proliferation rate of callus tissue being a critical factor contributing to the extended transformation period.This study aims to investigate the optimal light conditions for upland cotton callus growing.The establishment of this research endeavor is poised to accelerate callus proliferation and ultimately provide a technical foundation for shortening the period of cotton genetic transformation.【Method】The hypocotyls of the upland cotton line WC were used as explants to induce callus tissues under four different lights:red,blue,red-blue(1:1),and white light(CK).The study aimed to investigate the varied effects of the different lights on callus induction and proliferation,determine the optimal light condition by comparing with the callus proliferation rate,morphological characteristics et al.under different light treatments.【Result】Different light treatments had a significant impact on the callus induction and growth.The red-blue light treatment exhibited a most positive effect on both callus inducing and proliferation,which observed on the 7th and 15th day.The fresh weight of callus at 7 d under the red-blue light(0.39 g)was the heaviest,followed by the blue light(0.34 g),then the white light(0.24 g)and the red light(0.23 g).The same fresh weight order of the callus was observed at 15 d,with the treatment of the red-blue light(1.15 g)>blue light(0.98 g)>white light(0.69 g)>red light(0.51 g).The callus weight under red-blue light was 1.65 times heavier than the control,increased by 16.5%and 125.5%compared to the treatment of only blue or red light,respectively.In line with this,the callus proliferation rate in the second week under red-blue light was as high as 14.67%,which is twice to that of the red-light treatment(7.17%).The expression level of those genes promoted to cell proliferation and somatic embryo regeneration was consistent with the phenotype,the highest level was under the red-blue light treatment.Furthermore,the activity of the catalase(CAT)was significantly increased under the red-blue light treatment,while the content of reactive oxygen species(ROS)was lower than control.【Conclusion】Employing different light conditions could result in the varied proliferation rate of cotton callus.The optimal light treatment is by red-blue light in a 1﹕1 ratio,followed by blue light and then white light.However,employing red light only does not favor the callus growth.The treatment of red-blue light(1﹕1)induces the expression of phytochromes and cryptochromes in the callus,increases the expression level of those genes promoted to callus growth.It also enhances the activity of catalase,reduces the content of ROS,and finally promotes the callus proliferation.