摘要
[目的]从中草药马兜铃中获得酪氨酸脱羧酶基因(tyrDC)互补DNA(cDNA)序列并进行同源性分析,以进一步达 到敲除tyrDC,减轻该药肾毒性的目的。[方法]参照已知的植物酪氨酸脱羧酶的保守性氨基酸序列设计简并引物,并通过 反转录聚合酶链反应(RT-PCR)从马兜铃中扩增出cDNA片段。将扩增的cDNA序列克隆并测序,用以设计特异引物,并 通过cDNA末端快速扩增(Rapid Amplification of cDNA Ends,RACE)从马兜铃中扩增出全长tyrDC cDNA。[结果]该克隆的 tyrDC cDNA的总长度为1 678 bp,包括一个编码516个氨基酸的1 551bp开读框(ORF)和一段127 bp的下游非翻译区 (3'UTR)。序列比较结果表明,由马兜铃tyrDC核苷酸序列推导的氨基酸序列分别与已发布的罂粟酪氨酸脱羧酶和唐松草酪 氨酸／多巴脱羧酶享有76％和79％的同源性。[结论]从马兜铃中扩增得到tyrDC cDNA全序列,且对酪氨酸脱羧酶的同源检 索显示,不同植物的酪氨酸脱羧酶之间都存在普遍的序列相似性。为去除马兜铃肾毒性奠定了基础。
[Objective] To obtain the tyrosine decarboxylase gene (tyrDC) from Aristolochia contorta Bge.and to assay its cDNA sequence and homologous analysis, thus to remove its nephrotoxieity. [Methods] The primers designed by referring to the conservative amino acid sequences of known plant tyrDC were used to amplify a fragment of cDNA from Aristolochia contorta Bge.by reverse transcription-polymerase chain reaction (RT-PCR). The amplified cDNA sequence was cloned and sequenced to design a pair of specific primers and to amplify a full-length tyrDC cDNA from Aristolochia contorta Bge.by rapid amplification of cDNA ends (RACE). [Results] The length of cloned tyrDC cDNA is 1678 base pairs (bp), which comprises an open reading frame ( ORF) of 1551 bp encoding 516 amino acids and a downstream untranslated region (3'UTR) of 127 bp. The results of sequence comparison indicated that the amino acid sequence deduced from the nucleotide sequence of tyrDC from Aristolochia contorta Bge.shares 76% homology with issued tyrosine decarboxylase of Papaver somniferum L. and 79% homology with tyrosine /DOPA decarboxylase from Thalictrum flavum subsp. glaucum. [Conclusion] The full-length tyrDC cDNA has been amplified from Aristolochia contorta Bge. and the homologous retrieve of tyrosine decarboxylase reveals an extensive sequence similarity among tyrosine decarboxylases of different plants. This will provide evidence for the romoval of nephrotoxicity of Aristolochia contorta Bge. .
出处
《广州中医药大学学报》
CAS
2005年第2期152-156,159,共6页
Journal of Guangzhou University of Traditional Chinese Medicine
基金
国家自然科学基金资助项目(编号:30271591)
关键词
马兜铃/化学
酪氨酸脱羧酶
基因
结构
植物
基因扩增
ARISTOLOCHIA CONTORTA BGE. /chemistry
TYROSINE DECARBOXYLASE
GENES, STRUCTURAL, PLANT
GENE AMPLIFICATION
