PMID: 29750492Feb 4, 2017Paper

Isolation, characterization and real-time RT-PCR for relative quantification of gene expression in H+-ATPase-defective mutants from Lactobacillus plantarum

Wei sheng wu xue bao = Acta microbiologica Sinica
Xiang ZhangYuru You

Abstract

The aim of this study was to isolate Lactobacillus plantarum acid-sensitive mutants with lower H+-ATPase activity, and to study the mechanism of H+-ATPase regulation in Lactobacillus plantarum. We used neomycin to isolate acid-sensitive mutants of L. plantarum, and measured H+-ATPase activity and lactic acid production of wild-type and mutants. Genomic DNA was extracted from the wild-type ZUST and two mutants (ZUST-1, ZUST-2), and used as PCR templates. H+-ATPase genes of the strain were amplified, and the PCR products were sequenced. Sequence similarity of H+-ATPase was analyzed. Real-time RT-PCR was used to evaluate the relative quantification of the H+-ATPase genes expression. The growth of the mutants was characterized in MRS broth, which revealed that their cell biomass and acid production were lower than that of the wild-type. H+-ATPase activity of the mutants ZUST-1 and ZUST-2 was 10.1% and 28.8% lower than that of the wild-type. Results showed that atpA gene of the mutants ZUST-1 and ZUST-2 existed 22 mutations by alignment of the wild-type sequence, and atpC gene of ZUST-2 existed 6 mutations. Mutants ZUST-1 and ZUST-2 atpA gene expression were 41.1% and 35.7% lower than that of the wild-type in exponential phase, 43.6...Continue Reading

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