• 中文核心期刊
  • CSCD来源期刊
  • 中国科技核心期刊
  • CA、CABI、ZR收录期刊

普城沙雷氏菌ACCC 02146产灵菌红素调控基因的鉴定

Prodigiosin-producing Genes in Serratia plymuthica ACCC 02146

  • 摘要:
      目的  鉴定影响普城沙雷氏菌ACCC 02146产灵菌红素能力的基因,构建ACCC 02146的转座子突变体文库,为进一步研究普城沙雷氏菌ACCC 02146灵菌红素合成机制奠定基础。
      方法  采用平板划线法从菌种保藏中心购买菌株和实验室保藏菌株中获得15株产灵菌红素的菌株。采用16S rRNA基因测序法鉴定各菌株,邻接建树将其分类,并比较不同类别菌株灵菌红素合成基因簇启动子序列差异,观察各菌株产色能力。对16S rDNA序列和灵菌红素合成基因簇启动子序列均有异于其他菌株的普城沙雷氏菌ACCC 02146合成灵菌红素的调控基因展开研究。构建ACCC 02146的转座子突变体文库,筛选出产色能力明显改变的克隆子并鉴定出对应的转座子插入突变基因。
      结果  该突变体文库中有74个突变体表现出产灵菌红素能力的变化。其中25个突变体转座子插入发生在pigApigBpigCpigDpigH 等5个灵菌红素合成簇基因上,49个突变体转座子插入基因为灵菌红素合成基因簇之外的基因。在鉴定到的产色能力改变的突变体中,麦芽糖O-乙酰基转移酶基因突变菌株有6个,二氢乳清酸脱氢酶基因突变菌株有4个,MarR家族转录因子SlyA基因突变体有3个,winged helix家族的双组分转录调控因子RstA基因突变体有3个,H-醌氧化还原酶亚基I基因突变菌株有3个,NADH-醌氧化还原酶G链基因突变菌株有3个,肽基脯氨酰异构酶B基因突变菌株有3个,其他突变基因对应的克隆子数量为1~2个。
      结论  在沙雷氏菌中,除了灵菌红素合成簇基因调控灵菌红素合成,推测灵菌红素合成簇外编码相关酶、转录调控因子和一些结构蛋白的基因通过直接或间接途径在不同程度上调控了灵菌红素的合成。

     

    Abstract:
      Objective   Genes relating to prodigiosin synthesis in Serratia plymuthica ACCC 02146 were identified, and a transposon mutant library on the strain constructed.
      Methods  Fifteen prodigiosin-producing microbes were obtained from a conservation center and a laboratory. After identification by 16S rRNA gene sequencing, they were classified according to the neighbor-joining trees. Promoter sequences of prodigiosin synthesis gene were analyzed, and color producing capacity of the individual strains evaluated. Selected strain was cloned and further studied to establish a library on the transposon mutants.
      Results   Differed from other strains in terms of 16S rDNA and promoter sequences of the prodigiosin biosynthesis gene clusters, S.plymuthica ACCC 02146 was selected to clone the candidate gene for further investigation. A transposon mutant library was constructed subsequently. In the library, of 74 mutants showing significant variations in prodigiosin-producing ability, 25 had the insertions in pigA, pigB, pigC, pigD, and pigH, while 49 in the genes outside the cluster. On color formation, 6 strains with the mutation on maltose o-acetyltransferase gene, 4 on dihydroorotate dehydrogenase gene, 3 on MarR family of transcription factor SlyA genes, 3 on two-component transcriptional regulator RstA of the winged helix family, 3 on NAD(P)H-quinone oxidoreductase subunit I gene, 3 on NADH-quinone oxidoreductase, chain G gene, 3 on peptidylprolyl isomerase B gene, and one to two on other genes were found possibly related to significant alterations on the prodigiosin production as well.
      Conclusion  Aside from the identified specific clusters of prodigiosin synthesis-associated genes, additional factors in the forms of enzymes, transcriptional regulators, and/or structural proteins were now speculated to also directly or indirectly contribute in varying degrees to the prodigiosin synthesis in Serratia sp.

     

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