NEXTflex® 16S V5 – V6 Amplicon-Seq Kit for Illumina® Library Prep

Validated library prep for bacterial profiling of plant samples using Illumina® sequencing platforms

 

Catalog# Product Name Quantity US List Price
NOVA-4205-01
NEXTflex® 16S V5 – V6 Amplicon-Seq Kit (4 Barcodes)
8 rxns $180 Buy Now
NOVA-4205-02
NEXTflex® 16S V5 – V6 Amplicon-Seq Kit (12 Barcodes) 24 rxns $423 Buy Now
NOVA-4205-03
NEXTflex® 16S V5 – V6 Amplicon-Seq Kit (48 Barcodes) 96 rxns $1,524 Buy Now
NOVA-4205-04
NEXTflex® 16S V5 – V6 Amplicon-Seq Kit (Barcodes 1- 96)
192 rxns $2,895 Buy Now
NOVA-4205-05
NEXTflex® 16S V5 – V6 Amplicon-Seq Kit (Barcodes 97 - 192)
192 rxns $2,895 Buy Now
NOVA-4205-06
NEXTflex® 16S V5 – V6 Amplicon-Seq Kit (Barcodes 193 - 288)
192 rxns $2,895 Buy Now
NOVA-4205-07
NEXTflex® 16S V5 – V6 Amplicon-Seq Kit (Barcodes 289 - 384) 192 rxns
$2,895 Buy Now

NEXTflex™ 16S V5-V6 Amplicon-Seq Kit
  • Fast and easy library prep protocol for 16S profiling of bacteria in plant samples
  • Low input - Only 1 ng of input required
  • Multiplex up to 384 libraries
  • Custom sequencing primers are not required
  • Automation protocols are now available for the PerkinElmer® Sciclone® NGS and NGSx Workstation to automate your 16S sequencing
  • Functionally validated with Illumina® sequencing platforms

The NEXTflex™ 16S V5 – V6 Amplicon-Seq Library Prep Kit is designed for bacterial profiling of plant samples using degenerate primers that target the fifth and sixth variable regions of the 16S rRNA geneBacterial profiling in plant samples can be difficult because of the homology between bacterial 16S rRNA and chloroplast 16S rRNA. The use of primers binding to 16S hypervariable regions V 1 – V4, which are traditionally used for bacterial profiling, amplify both the bacterial and plant 16S rRNA genes, skewing results. NEXTflex™ 16S V5 – V6 Amplicon-Seq libraries are compatible with paired-end sequencing on the Illumina® MiSeq® sequencing platform and are ideal for monitoring population fluctuations of microbial communities.

Difference in Representation between 16S V4 and 16S V5-V6 in the Presence of Plant Contamination

Difference in Representation between 16S V4 and 16S V5-V6 in the Presence of Plant Contamination

The difference between the 0% and 90% plant-contaminated samples was calculated for each species and each primer set.  The 16S V4 amplicons show significant variability in the representation of bacterial community members with increasing plant DNA contamination.  In contrast, the 16S V5-V6 amplicons show very little sensitivity to the addition of up to 90% of plant DNA.


Optimized Protocol Offers Lower PCR Bias and Fewer Off-target Reads

The NEXTflex 16S V5 – V6 Amplicon-Seq Kit incorporates a second PCR step in the protocol for the addition of the sample-specific index, which reduces off-target reads encountered during amplicon sequencing.


Low Input Requirements for 16S rRNA Sequencing

As little as 1 ng of genomic DNA can be used to generate libraries using the NEXTflex 16S V5 – V6 Amplicon-Seq Library Prep Kit.


384 Barcodes Available for Cost-effective 16S V5 – V6 rRNA Sequencing

The NEXTflex 16S V5 – V6 Amplicon-Seq Library Prep Kit can be used for multiplexing up to 384 samples, to greatly reduce the cost of sequencing. The barcodes are offered in sets of 4, 12, 48, or 96 barcodes and supplied with 2 reactions worth of each barcode.


For Research Use Only. Not for use in diagnostic procedures.

Kit Specs

The NEXTflex 16S V5 – V6 Amplicon-Seq Kit contains enough material to prepare 8, 24, 96, or 192 amplicon-seq libraries from bacterial genomic DNA for sequencing using the Illumina® MiSeq® platform. The shelf life of all reagents is 12 months when stored properly. All components can be safely stored at -20°C. This kit is shipped on dry ice.

NEXTflex 16S V5 – V6 Amplicon-Seq Kit Protocol

NEXTflex 16S V5-V6 Amplicon-Seq Kit Protocol

 

Kit Contents

NEXTflex™ PCR Master Mix

NEXTflex™ 16S V5-V6 PCR I Primer Mix

NEXTflex™ PCR II Barcoded Primer Mix

Resuspension Buffer

Nuclease-free Water

 

Required Materials Not Provided

1 ng - 50 ng high-quality genomic DNA in up to 36 µL nuclease-free water for each library

96 well PCR Plate Non-skirted (Phenix Research, Cat # MPS-499) or similar

Adhesive PCR Plate Seal (Bio-Rad®, Cat # MSB1001)

Agencourt® AMPure® XP 5 mL (Beckman Coulter® Genomics, Cat # A63880)

Magnetic Stand - 96 (Thermo Fisher Scientific®, Cat # AM10027) or similar

Thermocycler

2, 10, 20, 200 and 1000 µL pipettes / multichannel pipettes

Nuclease-free barrier pipette tips

Vortex

80% Ethanol, freshly prepared (room temperature)

Sequences of NEXTflex 16S V5 – V6 Amplicon-Seq Barcoded Primers Indexes - Excel / PDF

Instructions for installing NEXTflex Barcode Indices in Illumina Experiment Manager