过表达FaSAMDC基因提高黑麦草属植物的抗旱性和耐热性

主要探讨了在黑麦草属植物中导入高羊茅S-腺苷甲硫氨酸脱羧酶基因(FaSAMDC)对转基因植株抗旱耐热性的提高效果,为培育黑麦草抗旱耐热新品种提供种质新材料。选择黑麦草成熟种子为外植体,采用贵州省草业研究所分子生物学实验室构建的过表达载体以及黑麦草胚性愈伤组织高频植株再生和农杆菌介导的遗传转化体系,将克隆自高羊茅的FaSAMDC基因转入贵州地区主栽品种多花黑麦草特高和多年生黑麦草四季的基因组内,经抗性筛选、PCR检测和Southern杂交分析验证,获得45株特高与44株四季的转基因植株,转化频率分别为2.93%和2.28%。抗旱耐热试验表明,在30 ℃高温和中度干旱胁迫条件下,特高转基因株系的根冠比比对照组培苗高出7.08%,叶片相对含水量(RWC)比对照高出13.12%,RWC降低幅度比对照少5.49个百分点;四季转基因株系的根冠比比对照组培苗高出6.54%,RWC比对照高出12.54%,RWC降低幅度比对照少6.50个百分点,差异均达到显著水平(P<0.05),证明转入FaSAMDC基因的黑麦草阳性株系的抗旱耐热能力得到了明显提高。形态与生长特征观测结果显示,与非转基因组培苗相比,转基因株系的叶长、株高、主茎节数减小,叶宽、单株分蘖数增加,叶色变深,植株形状趋向于叶片丛生的紧凑型,推测导入的FaSAMDC基因参与了基因表达、细胞分裂等生理功能的调节。

Enhancing drought and heat tolerance in Lolium spp.through overexpression of the FaSAMDC gene

To obtain new ryegrass germplasm with enhanced drought and heat tolerance a genetic transformation experiment using plants from the genus Lolium was conducted.By using mature seeds of ryegrass as explants and adoption of the overexpression vector,tissue culture and regeneration system of ryegrass,and genetic transformation mediated by Agrobacterium tumefaciens,the S-adenosylmethionine decarboxylase gene cloned from Festuca arundinacea (FaSAMDC) was introduced into the genome of 'Tetragold' annual ryegrass and 'Four Seasons' perennial ryegrass.Eighty-nine transgenic plants (45 Tetragold and 44 Four Seasons) were regenerated from independent resistant calli,which were confirmed by antibiotic resistance screening,PCR assay and southern hybridization analysis;the transgenic frequency was 2.93％ and 2.28％,respectively.Drought and heat resistant tests under high temperature (30 ℃) and moderate drought stress showed that in Tetragold,the root to shoot ratio was 7.1％ higher,leaf relative water content (RWC) 13.1％ higher and the RWC descent scope 5.5％ lower than the controls;for transgenic Four Seasons plants the root to shoot ratio was 6.5％ higher,RWC 12.5％ higher and the RWC descent scope 6.5％ lower than the control;differences between transgenic plants and controls were all statistically significant (P＜0.05),indicating that drought and heat resistance in transgenic plants was significantly improved.Observations of morphological and growth characteristics showed that,compared with non-transgenic plants,leaf length,plant height and main stalk pitch number of transgenic plants were lower but leaf width and tiller number per plant were higher.Leaves of transgenic plants were darker in colour and plants tended to become compact with tufted leaves suggesting that the inserted FaSAMDC gene was involved in the regulation of physiological function,such as gene expression and cell division.The transgenic material developed in the study could be used as foundation for the cultivation of new ryegrass varieties with enhanced drought and heat resistance.