大庆湿地可培养噬藻体的分离及其相关基因的系统进化分析

Isolation and phylogenetic analysis of related genes of cultivable cyanophages in Daqing Wetland

摘要:

[目的] 揭示大庆湿地可培养蓝藻噬菌体遗传基因多样性,分析其系统进化地位,为噬藻体生态学研究提供数据支持。[方法] 以鱼腥藻(Anabaena PCC7120)为宿主,采用液体富集和双层平板法分离大庆湿地水体中可培养的噬藻体,提取噬藻体混合液的DNA, PCR扩增噬藻体编码衣壳组装蛋白的g20基因和编码T7型短尾病毒的核糖体聚合酶的pol基因,克隆测序,构建系统进化树,明确可培养噬藻体相关基因的系统进化地位。[结果] 克隆测序获得1条g20基因序列,4条pol基因序列。系统进化分析表明,获得g20序列隶属于可培养噬藻体类群(Cluster δ)中。而3条pol基因与我国吉林碱性稻田水体噬藻体类群(PG-Pol-I和PG-Pol-II)更相近,另一条pol序列形成独立的进化分枝。[结论] 这是首次调查大庆湿地水体侵染鱼腥藻的可培养噬藻体的g20和pol基因,初步确认以鱼腥藻(Anabaena PCC7120)为宿主的可培养噬藻体g20基因归属于Cluster δ中,而大庆湿地可培养噬藻体的pol基因与我国大安稻田水体pol基因相近。

关键词:大庆湿地;可培养噬藻体;鱼腥藻;系统进化

Abstract:

[Objective] To provide scientific data for studying the ecology of cyanophages isolated from Daqing wetland by analyzing their genetic diversity and phylogenetic positions. [Methods] Liquid enrichment culture and double-layer plate methods were used to isolate cultivable cyanophages by using host cyanobacteria Anabaena PCC7120, and the DNA of cultivable cyanophages was extracted. The biomarker genes of g20 encoding capsid assembly protein and pol encoding DNA polymerase in podoviruses were PCR amplified. The PCR products were cloned and sequenced. The sequences were constructed with references sequences into the phylogenetic trees to clarify the phylogenetic positions of cultivable cyanophage. [Results] One g20 sequence and four pol sequences were obtained. Phylogenetic analysis showed that the g20 sequence belongs to the cultivable cyanophage group (Cluster δ). Three pol sequences were closely related to cyanophage groups PG-Pol-I and PG-Pol-II that were observed in an alkaline paddy floodwater in Da'an, Jilin province, China; another pol sequence formed a unique clade. [Conclusion] The g20 gene from cultivable cyanophages infecting Anabaena PCC7120 belongs to the Cluster δ, and the pol genes are closely related to those of paddy floodwater in Da'an, China.

Key words:Daqing Wetland;cultivable cyanophage;Anabaena;phylogenetic analysis