根据在GenBank中登录的番茄、马铃薯等蔗糖磷酸合成酶基因的保守序列设计引物,采用RT-PCR方法从甜瓜花后25d的果实总RNA中扩增出目标cDNA片段,克隆到pMD18-T载体中。序列分析表明,该片段与番茄蔗糖磷酸合成酶氨基酸序列同源性为98.9%,GenBank中登记号为DQ355797。通过Northern blot检测其在甜瓜果实不同发育时期的表达变化,结果表明该基因在甜瓜果实花后25d开始表达,随着果实的成熟,表达量升高。
PCR primers were designed based on the conserved domain of some sucrose phosphate synthase genes in GenBank. The target cDNA fragment was amplified from the total RNA isolated from melon fruit of 25 days after pollination with RT-PCR and then was cloned into pMD18-T vector. The sequence showed that the amino acid sequence encoded by this fragment showed a 98.9% similarity to that of the corresponding region of sucrose phosphate synthase gene in tomato. The accession number of this gene in GenBank was DQ355797. Northern blot was performed to analyze the expression pattern of sucrose phosphate synthase in different development stages of melon fruit. The result showed that sucrose phosphate synthase began to express in fruit 25 days after pollination and the expression level of this gene increased with melon fruit mature.