Identification of AlmA genes involved in long-chain alkane degradation by Alcanivorax hongdengensis A-11-3
【目的】研究海洋烷烃降解菌新种模式菌株AlCAnIVOrAX HOngdEngEnSIS A-11-3降解长链烷烃的分子机制。【方法】PCr克隆编码黄素结合单加氧酶的基因序列,利用生物信息学软件对序列进行分析,运用rT-PCr和实时荧光定量PCr技术分析基因在不同烷烃诱导下的表达水平。【结果】从菌株A-11-3中克隆获得了两个黄素结合单加氧酶基因片段(AlMA1和AlMA2)。它们编码的氨基酸序列与菌株ACInETObACTEr SP.dSM17874的AlMA同源性分别为58.6%和53.2%。实时荧光定量PCr分析表明,AlMA1基因只在长链烷烃(C28-C32)的诱导下上调表达,而AlMA2基因中能在更宽范围的长链烷烃(C24-C34)和支链烷烃诱导下上调表达。两者均在C9-C22的烷烃诱导下没有上调表达。【结论】黄素结合单加氧酶可能是A-11-3降解长链烷烃和支链烷烃的关键酶。[Objective] The aim of this study is to identify of genes involved in long-chain (LC) alkane degradation in Alcanivorax hongdengensis A-11-3.[Methods]PCR was applied to obtain Flavin-binding monooxygenase genes,then quantitative real-time polymerase chain reaction ( Q-RT-PCR) and RT-PCR were applied to analyze gene expression in response to different LC-alkanes and pristane.[Results] Two homologues,almA1and almA2,were obtained.They showed 58.6% and 53.2% similarities with almA of Acinetobacter sp.Strain DSM 17874,respectively,at amino acid level.Enhanced expression of almA1 genes was observed when strain A-11-3 grew with long chain alkanes (C28 to C 32),in sodium acetate medium.However,the induction expression was not observed in the case of C9-C22 alkanes.Similarly,almA2 was induced by long chain alkanes (C24 to C 34 ).In addition,it was also induced by the branched alkane pristane.[Conclusion]AlmA genes were mostly responsible for the degradation of long-chain alkanes and pristane in strain A-113.