中文摘要：

【目的】克隆桃果实中的磷脂酶Dα基因,并对其进行序列分析和低温表达模式分析。【方法】采用电子克隆与RT-PCR相结合的方法分离桃果实PLDα基因cDNA全长序列;半定量RT-PCR和Northern杂交检测目的基因在桃果实低温贮藏过程中的表达情况。【结果】成功获得全长为2859bp的桃果实PLDα基因序列,该序列具有完整的开放阅读框架（ORF,172～2604bp）,编码810个氨基酸。对ORF进行的RT-PCR验证结果与电子克隆序列一致。序列分析显示,桃果实PLDα基因编码的氨基酸序列与草莓、番茄、葡萄等物种的磷脂酶Dα氨基酸序列之间具有高度的保守性,含有N端C2结构域及两个保守的活性中心。半定量RT-PCR和Northern杂交分析显示,桃PLDα基因在低温贮藏过程中被诱导表达。【结论】本研究首次通过电子克隆的方法获得了桃果实PLDα基因的全长序列,揭示了其序列特征,并初步明确了其在桃低温贮藏期间的表达模式。结果证明基于EST数据库的电子克隆已经是获取植物新基因的有效途径,同时为更深入研究桃果实的耐冷性及低温贮藏方法奠定了基础。

英文摘要：

[Objective] The aim of this study was to isolate and clone new gene coding for phospholipase Dα from Prunus persica. [Method] In silico cloning was used to isolate cDNA sequence ofPLDa, RT-PCR and Northern blot analysis were used to test the sequence and to analyze its expression during storage at 4℃. [Result] The obtained sequence contained a complete ORF, from 172 bp to 2 604 bp, encoded 810 amino acids. Compared with other plants, such as tomato, strawberry and grape etc., it was found that the amino acids sequence encoded by phospholipase Dot gene was almost conserved. Semi-quantitative RT-PCR and Northern blot analysis indicated that PLDa could be induced by low temperature. [Conclusion] Full length PLDa gene was first isolated and characterized from peach fruit, suggesting it is an efficient technique for cloning novel genes by searching EST database with homologous gene of relative species. Transcription profile of PLDa mRNA has provided a scientific basis for further research on the relationship between PLDa and low temperature.