16S ribosomal RNA (rRNA) sequencing is a common amplicon sequencing method used to identify and compare bacteria present within a given sample. 16S rRNA gene sequencing is a well-established method for studying phylogeny and taxonomy of samples from complex microbiomes or environments that are difficult or impossible to study.
- Submit 1-5 µg of RNA-free genomic DNA (NGS grade) of high molecular weight, Nanodrop A260/280 ratio >1.8; A260/230=2.0-2.2; and at a concentration of ~50 ng/µl
- Or soil, plant/ human microbiome, water or sludge samplesshipped in cool packs to eurofins facility.
- RNA free NGS grade gDNA isolation will be carried out using commercially available DNA isolation kit.
- Further quality checking will be carried out using agarose electrophoresis and NanoDrop. Quantification will be carried out using Qubit 3.0 fluorometer.
- The amplicon libraries will be prepared using 16S rDNA gene specific primers and indexing using Nextera XT Index Kit.
- Library validation will be carried out using High Sensitivity D1000 Screen tape using Agilent Tape station kit.
- Prepared library will be sequenced using 2 x 300 bp paired end sequencing using latest sequencing chemistry on illumina MiSeq.
- Delivery of data up to ~5, 00,000 paired end reads per sample.
- Sequence data (base call files or bcl files) generated from the sequencer are de-multiplexed if required and converted to FASTQ files.
- Raw data will be available for download as a compressed archive of FASTQ files for each sample.
- Raw data can be post processed upon request.
- 5-7 weeks after arrival of your samples and all necessary information. It depends upon data size, scope, technology selected, number of samples and complexity of the project. The TAT is offered expecting no biological or technical difficulties for processing of all project samples.