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目的克隆刺五加亚精胺合成酶(spermidine synthase,SPDS)基因,并分析内生真菌对其表达的影响。方法采用cDNA末端快速扩增(rapid amplification of cDNA ends,RACE)技术克隆刺五加SPDS基因全长cDNA序列。运用生物信息学方法对该基因进行分析。RT-PCR法检测内生真菌菌株P116-1a、P116-1b、P109-4和P312-1对SPDS基因表达的影响。结果刺五加SPDS基因的cDNA全长为1 541 bp,开放阅读框长1 002 bp,编码333个氨基酸的蛋白,包含SPDS家族的基本结构和标志性序列。RT-PCR结果显示,内生真菌可显著提高刺五加SPDS基因的表达量(P<0.05),最大表达量出现在菌株P116-1b回接90 d时,是对照的2.06倍。结论首次克隆了刺五加SPDS基因的cDNA全长序列,并证实内生真菌可显著提高刺五加SPDS基因的表达,为阐明内生真菌提高刺五加三萜皂苷量的机制及刺五加的抗逆性改良奠定了基础。
Objective To clone the gene of spermidine synthase (SPDS) and analyze the effect of endophytic fungi on its expression. Methods The full length cDNA sequence of Acanthopanax senticosus SPDS was cloned by rapid amplification of cDNA ends (RACE). This gene was analyzed using bioinformatics methods. The effects of endophytic fungi strains P116-1a, P116-1b, P109-4 and P312-1 on the expression of SPDS gene were detected by RT-PCR. Results The full-length cDNA of SPDS gene of Acanthopanax senticosus was 1 541 bp in length with an open reading frame of 1002 bp encoding a protein of 333 amino acids, including the basic structure and landmark sequence of the SPDS family. The results of RT-PCR showed that the endophytic fungi could significantly increase the expression of SPDS gene of Acanthopanax senticosus (P <0.05), and the maximum expression level appeared 2.06 times of the control when the strain P116-1b was connected 90 days later. Conclusion The full-length cDNA of SPDS gene of Acanthopanax senticosus was cloned for the first time. It was confirmed that endophytic fungi could significantly increase the expression of SPDS gene of Acanthopanax senticosus. To elucidate the mechanism of increasing the amount of acanthopanax senticosus saponins by endophytic fungi, The improvement of stress resistance laid the foundation.