采用IEF/SDS-PAGE双向电泳技术,对‘立冬本’龙眼合子胚发育不同时期的蛋白质进行分离,通过MALDI-TOF/TOF鉴定到其中一个上调差异表达的蛋白为黄烷酮-3-羟化酶(flavanone 3-hydroxylase,F3H)。以‘立冬本’龙眼胚胎cDNA为模板,采用RT—PCR和RACE技术,获得F3H基因cDNA1404bp全长序列。序列分析表明,F3H基因共编码365个氨基酸,其cDNA序列与柑橘、苹果、棉花等F3H基因的同源性均高达80%以上,该基因在GenBank中登录号为EF468104。
The protein of 'Lidongben' longan zygotic separated and compared by IEF/SDS-PAGE technique embryos during different development stages were Flavanone 3-hydroxylase was one of differential proteins identified by MALDI-TOF/TOF. Then F3H gene cDNA sequence was cloned from longan zygotic embryos using RT-PCR and RACE technique. A 1 404 bp full-length cDNA sequence was obtained. Analysis of F3H gene cDNA indicated that it encoded a peptide containing 365 amino acids. The nucleotide sequence comparison with the F3H gene of Citrus sinensis, Gossypium hirsutum and Pyrus communi showed that identity was all higher than 80%. The sequence was accepted and released by GenBank (Accession number: EF468104).