A NOVEL METHOD FOR ASSESSMENT OF 16S RRNA GENE COPY NUMBER IN BACTERIAL GENOMES BY PULSED-FIELD GEL ELECTROPHORESIS AND PCR AMPLIFICATION

摘要

16S rRNA gene (rrn) copy number in bacterial genomes is indicative of ecological strategies of bacteria and is critical for quantification of bacterial abundance in mixed populations using rrn-based approaches. For accurate assessment of rrn copies, a novel technical strategy by means of pulsed-field gel electrophoresis and polymerase chain reaction amplification analysis was introduced. Experimental and in silico analysis on a test bacterial culture Caulobacter crescentus proved it to be simple, effective, accurate and a good alternative to traditional time-consuming methods. This method can be used for routine determination of gene copy number in most bacteria whose full genome sequences are not available. Moreover, the pulsed-field gel electrophoresis bands containing a target gene fragment can be determined and therefore constructing an expected fragments oriented genomic library is possible.