草甘膦抗性菌株的分离鉴定及其抗性基因的克隆

从草甘膦污染土壤中分离获得一株耐受400 mmol/L草甘膦的菌株S1536,该菌株在草甘膦浓度为100~400 mmol/L之间生长迅速,最适pH为6.0,最适生长温度为30℃,具有氨苄青霉素抗性。以S1536基因组DNA为模板,通过通用引物进行扩增、测序获得其16S rDNA序列,在Gen Bank登录号为MG519831,在NCBI中经BLAST比对与泛菌属(Pantoea sp.)同源性达到99%。用ClustalW对S1536与泛菌属各个种模式菌的16S rDNA进行多重序列对比,该菌株与Pantoea rodasii同源性最为接近,达到99.2%,故将菌株S1536命名为Pantoea rodasii S1536。该菌株具有较高草甘膦抗性,对温度和pH适应性强,是优良的耐草甘膦菌株,通过全基因组测序及基因注释,获得抗除草剂基因aroA序列,为下一步明确基因功能,挖掘Pantoea rodasii S1536高抗草甘膦的分子机制奠定基础。

Isolation,Identification and Its Resistant Gene Cloning of a New Glyphosate-resistant Strain

A 400 mmol/L glyphosate-resistant strain S1536 was isolated from the glyphosate polluted soil. The strain grew well in the medium with glyphosate concentration between 100～400 mmol/L. The optimal pH was 6.0, optimal growth temperature was 30℃, and had resistance to ampicillin. Taking S1536 genomic DNA as a template, this experiment carried out amplification and sequencing and gained 16S rDNA sequence, which registration number was MG519831 in GenBank. Compared with the published nucleotide sequence of 16S rDNA in NCBI (National Center for Biotechnology Information), S1536 showed 99% homology with Pantoea sp. Multiple sequence comparison between S1536 and the model bacteria of Pantoeasp of 16S rDNA was conducted by ClustalW. This strain showed high identity with Pantoea rodasii, reaching 99.2%, so it was named as Pantoea rodasii S1536. The strain possessed higher glyphosate resistance and good adaptability to temperature and pH. It was a superior glyphosate tolerant strain. aroA sequence resistant to herbicide was obtained through whole genome sequencing and gene annotation, which provided a solid foundation for further identification of gene function and exploitation of the molecular mechanism of high glyphosate resistance of Pantoea rodasii S1536.