甘蔗△1-吡咯啉-5-羧酸合成酶基因转化烟草的抗旱性分析
Drought resistance of transgenic tobacco with sugarcane △1-pyrroline-5-carboxylate synthase (SoP5CS) gene

为了分析甘蔗P5CS（sugrcane △1-pyrroline-5-carboxylate synthetase gene,SoP5CS）基因的抗旱性能，利用农杆菌介导的叶盘转化法对烟草进行遗传转化，对拷贝数较低的4个转基因株系（T1、T4、T6和T9）进行抗旱性分析。结果显示：在干旱胁迫下，T9株系中SoP5CS及其编码的蛋白的相对表达量都是最低的，T1、T4、T6中SoP5CS的相对表达量分别是T9的9.02、3.93和8.51倍，SoP5CS蛋白则分别是T9的2.22、3.91和1.36倍；进一步分析发现T1和T6叶片脯氨酸和ABA含量相对较高，且4个株系均显著高于对照；MDA含量则是对照显著高于转基因植株，其中T1的MDA累积量最低；P5CR和P5CS活性在对照和转基因植株间表现不一，在T1和T4中，2种酶活性均与对照相当，T6和T9则是显著高于对照；对于CAT活性，表现为T6最高，T1、T4、T9也显著高于对照；对于SOD活性，转基因株系均显著比对照高，其中T4和T6的相对较高；转基因株系的叶片相对含水量明显高于对照，叶绿素减少量则低于对照。综合分析发现,T6株系的抗旱性表现最好，T1株系较好，表明SoP5CS基因确实具有较强的抗旱功能。
In order to analyze the drought resistance of sugarcane △1-pyrroline-5-carboxylate synthase gene (SoP5CS),SoP5CS was transformed into tobacco using Agrobacterium-mediated leaf discs method. Four transgenic tobacco lines with low copy numbers were screened for drought stress experiment. The results showed that both the relative expression levels of SoP5CS and its encoded protein were the lowest in T9 transgenic tobacco line under drought stress.Compared with T9,the relative expression level of SoP5CS in T1,T4 and T6 lines were 9.02-fold,3.93-fold and 8.51-fold while the protein level of SoP5CS in T1,T4 and T6 lines was 2.22-fold,3.91-fold and 1.36-fold,respectively.Results of further analyses showed that the contents of proline and ABA in the four transgenic lines were all significantly higher than those in the control.The contents of proline and ABA in T1 and T6 lines were higher than those in the other two lines. The MDA content in the control was significantly higher than that of the transgenic plants.The accumulation of MDA was the lowest in T1 line. The P5CR and P5CS activities were different between the control and transgenic plants.The activities of both enzymes in T1 and T4 lines were nearly the same as those in the control,significantly higher than those in T6 and T9 lines. The CAT activity was the highest in T6 line,which in the other transgenic lines significantly higher than that of the control. SOD activity in all the transgenic lines was significantly higher than that in the control,with which in T4 and T6 lines being relatively higher. The relative water content in the leaves of the transgenic lines was significantly higher than that in the control,while the chlorophyll reduction was lower. The result of RA comprehensive analysis showed that the drought resistance of T6 line was the best,followed by the T1 line,indicating that drought resistance of SoP5CS gene is strong.