V3-V4 regions of the 16S rRNA gene of Bacteria: Aedes albopictus larval habitat water
DNA extraction from larval habitat water samples was performed using a QIAamp Fast DNA stool mini kit. Briefly, a piece of membrane filter through which the water had been passed was extracted in optimized buffers in combination with inhibitEX buffer to remove PCR inhibitors, as recommended by the manufacturers. DNA concentration and purity were monitored using a NanoDrop Spectrophotometer and electrophoresis through 1% agarose gels. The next-generation sequencing analysis was based on the 16S rRNA gene of bacteria. The V3-V4 regions of the gene were amplified using universal region-specific primers 341F (5-CCT AYG GGR BGC ASC AG-3) and 806R (5-GGA CTA CNN GGG TAT CTA AT-3) tagged with sample-identifying barcodes. PCR products of the proper size were selected by 2% agarose gel electrophoresis. The DNA fragments were end-repaired and A-tailed then further ligated with Illumina adapters. The libraries were generated and sequenced on a paired-end Illumina platform.