低温胁迫对转SoTUA基因甘蔗生理生化特性的影响

Effects of low temperature stress on physiological and biochemical characteristics of SoTUA transgenic sugarcane

[目的]研究低温胁迫对转α-微管蛋白(TUA)基因甘蔗生理生化特性的影响,对甘蔗TUA基因(SoTUA)进行功能鉴定,为甘蔗抗寒品种选育提供参考依据。[方法]通过对pCAMBIA3300载体上的标记基因GFP进行PCR扩增及测序,鉴定转SoTUA基因阳性甘蔗品系。从中选择4个转基因品系(L1、L2、L3和L4),以野生型ROC22植株为对照(CK),对各处理进行低温(4 ℃)胁迫处理,在胁迫第0、4、7和12 d时分别测定不同处理甘蔗的生理生化指标。[结果]运用PCR筛选得到转SoTUA基因T1代甘蔗植株,对其中4个品系及CK的低温胁迫试验结果表明,在低温下各品系可溶性糖含量以不同模式变化,转基因各品系普遍高于CK;各品系中的可溶性蛋白在冷驯化前期强烈诱导,总体变化趋势为先升高后降低,转基因各品系普遍高于CK;各品系中丙二醛(MDA)含量增加,其中CK的增幅最大;过氧化物酶(POD)活性下降,除L2外,POD活性变化总体上呈先升高后降低的变化趋势;脯氨酸含量在低温胁迫过程中的变化趋势无明显规律。利用改进的极点排序法对各品系抗寒性进行评价,可知各品系抗寒性强弱排序为:L3>L1>L2>L4>CK。[结论]低温胁迫下4个转SoTUA基因甘蔗品系在生理生化指标上的表现均优于CK,即SoTUA基因在甘蔗体内的过量表达有利于甘蔗抗寒性能的提高。

【Objective】This study was conducted to investigate the effects of low temperature stress on physiological and biochemical characteristics of α-tubulin(TUA)transgenic sugarcane,and to identify the function of sugarcane TUA gene(SoTUA). It provided reference for sugarcane cold-resistant variety breeding.【Method】SoTU Atransgenic sugarcane lines were identified by PCR amplification and sequencing of GFP marker gene on vector pCAMBIA3300. Four transgenic lines(L1,L2,L3 and L4)were chosen for a low temperature(4 ℃)stress assay when the wild type of sugarcane variety ROC22 was used as control(CK). The changes of physiological and biochemical indexes were measured at 0,4,7,12 d after treatment.【Result】The SoTUA transgenic T1 sugarcane plants were screened by PCR,and the results of low temperature stress assay on four lines and the control showed that the soluble sugar content of each line varied in different patterns at low temperature,and the transgenic lines were generally higher than the CK. Soluble protein of all lines was strongly induced in the early stage of cold treatment,the overall change trend was first increased and then decreased,and the trans genic lines were generally higher than the CK. Malonaldehyde(MDA)content in each line increased,especially in the control. Peroxidase(POD)activity decreased,except for L2,POD activity generally increased and then decreased. The proline content change had no obvious regulations during the low temperature stress. The improved pole ordination method was used for the comprehensive evaluation of cold resistance. The results showed that the cold resistance of all the lines was ranked as L3>L1>L2>L4>CK.【Conclusion】The physiological and biochemical performances of the four SoTUA transgenic sugarcane lines were better than the CK under low temperature stress. Therefore,it can be concluded that the overexpression of SoTUA gene in sugarcane is conducive to the improvement of cold resistance.