山菠菜胆碱单加氧酶基因对棉花的遗传转化和耐盐性表达

胆碱单加氧酶（CMO）是渗透保护剂甜菜碱生物合成的关键酶之一。以棉花（Gossypium hirsutium L.）泗棉3号的下胚轴切段为外植体，利用农杆菌介导法将克隆自山菠菜（Atriplex hortensis）的AhCMO基因导入其中，通过组织培养胚状体发生途径获得转基因再生植株。以0.5%卡那霉素对再生苗筛选后，PCR检测抗卡那霉素棉苗确认阳性转化株，Southern和Northern杂交结果进一步证实外源基因已导入棉花并得到表达。在温室盆栽条件下，于2片真叶期对转AhCMO基因棉花及其转化受体泗棉3号施加0.5%的NaCl胁迫，15 d后发现其光合作用和植株生长被显著抑制，非转基因棉花泗棉3号的株高、鲜重和光合速率分别降低了57.6%、65.6%和69.9%，而转AhCMO基因的棉株分别降低了37.3%、54.6% 和47.9%。转AhCMO棉花所受盐害程度显著小于非转基因的棉株，说明AhCMO基因的导入和表达提高了转基因棉花的耐盐性。

Transformation of Cotton (Gossypium hirsutum L.) with AhCMO Gene and the Expression of Salinity Tolerance

Soil salinity is becoming a serious threat to global agriculture including cotton. Although cotton has been classified as a salt-tolerant crop, it is also seriously attacked by salinity especially during emergence and seedling growth. Levels of glycine betaine, an osmoprotectant accumulated in plants during abotic stresses, vary among cotton genotypes, and positively correlate with the degree of salt tolerance. Therefore, enhancing glycine betaine synthesis is one of the most promising ways to improve salt tolerance in cotton. Choline monooxygenase (CMO) catalyzes the committed step in the synthesis of glycine betaine. AhCMO, a gene cloned from Atriplex hortensis, was introduced into the cotton (Gossypium hirsutium L.) hypocotyl explants of SM3 via Agrobacterium mediation, and the transformed plants were regenerated through somatic embryogenesis in tissue culture. After selection with 0.5% kanamycin, the kanamycin-resistant regenerated plants were confirmed to be electropositive by polymerase chain reaction (PCR). Southern and Northern blotting analyses further indicated the introduction and the expression of AhCMO gene in transgenic cotton plants, respectively. At the two true-leaf stage, transgenic cotton seedlings were treated with 0.5% NaCl for 15 days under greenhouse conditions, plant height, fresh weight per plant and net photosynthetic rate were determined. The results showed that NaCl stress decreased plant height, fresh weight per plant and net photosynthetic rate by 37.3%, 54.6%, and 47.9% for transgenic plants, and by 57.6%, 65.6%, and 69.9% for non-transgenic SM3, compared with their corresponding NaCl-free controls, respectively. The fact that less injury of NaCl to transgenic plants than to non-transgenic plants, suggested that induction and expression of AhCMO considerably enhanced salinity tolerance of transgenic cotton plants. Transgenic cotton with improved tolerance against salt stress is of great agronomic value. However, it should be noted that the improvement in salt tolerance of the transgenic lines is still limited. An integration of the transgenic technology and the traditional breeding technique may further improve both salt-tolerance and other agronomic properties of cotton.