NEXTflex® Congenital Adrenal Hyperplasia Amplicon Panel

Validated panel for detection of the CYP21A2 gene with library prep reagents and 384 barcodes for Illumina® sequencing

Catalog# Product Name Quantity US List Price
4244-01
NEXTflex® Congenital Adrenal Hyperplasia Amplicon Panel for Illumina® platforms (8 Barcodes)
8 rxns $375 Buy Now
4244-02
NEXTflex® Congenital Adrenal Hyperplasia Amplicon Panel for Illumina® platforms (48 Barcodes)
48 rxns $2,100 Buy Now
4244-03
NEXTflex® Congenital Adrenal Hyperplasia Amplicon Panel for Illumina® platforms (Barcodes 1- 96)
96 rxns $3,888 Buy Now
4244-04
NEXTflex® Congenital Adrenal Hyperplasia Amplicon Panel for Illumina® platforms (Barcodes 97 - 192)
96 rxns $3,888 Buy Now
4244-05
NEXTflex® Congenital Adrenal Hyperplasia Amplicon Panel for Illumina® platforms (Barcodes 193 -288)
96 rxns $3,888
Buy Now
4244-06
NEXTflex® Congenital Adrenal Hyperplasia Amplicon Panel for Illumina® platforms (Barcodes 289 - 384) 96 rxns
$3,888
Buy Now

NEXTflex CAH Amplicon Panel
  • 100% coverage of the CYP21A2 exons and flanking intron-exon boundaries 
  • 100% uniformity of amplicons on 0.2x coverage
  • 384 unique barcodes available
  • Complete solution for targeted sequencing, including gene specific primers, PCR Master Mix, clean-up beads, and up to 384 barcodes
  • Low input - Only 20 ng DNA isolated from fresh or frozen samples required

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

Complete Coverage of the CYP21A2 Gene for Illumina® Sequencing

The NEXTflex® Congenital Adrenal Hyperplasia Amplicon Panel allows you to quickly target and sequence 10 exons of the CYP21A2 locus. This kit contains primer pairs and reagents needed to amplify these coding regions of the CYP21A2 gene from DNA isolated from fresh or frozen material, and prepare libraries for Illumina® sequencing.

Traditional amplicon sequencing of CYP21A2 can be confounded by pseudogene CYP21A1P, which has 99% homology with CYP21A2. In order to obtain ‘true’ CYP21A2 reads, the NEXTflex Congenital Adrenal Hyperplasia Amplicon Panel uses a two-step enrichment. In the first step, sample DNA is enriched for CYP21A2 DNA by long-range PCR. In the second step, ten canonical exons are targeted and amplified. During a gene conversion event, CYP21A2 and CYP21A1P can transfer pseudogene sequences into the CYP21A2 gene. To detect CYP21A2 mutations introduced by the pseudogene, enrichment primers are designed to amplify both CYP21A2 and CYP21A1P with the same efficiency.

While up to 384 barcodes are currently supplied with the NEXTflex Congenital Adrenal Hyperplasia Amplicon Panel, the panel has the capability of multiplexing up to 950 samples at 100x coverage for detection of germline mutations on a single Illumina® 2x300 MiSeq® lane. If you need the ability to multiplex more than 384 samples, contact nextgen@biooscientific.com to request a quote for a panel with additional barcodes.


A Complete Solution for Targeted CYP21A2 Sequencing

The NEXTflex Congenital Adrenal Hyperplasia Amplicon Panel is a complete, validated solution that targets 10 exons in the CYP21A2 locus using 20 PCR primer pairs. This kit requires only 20 ng of DNA isolated from fresh or frozen samples. The convenient predesigned panel allows researchers to focus on data generation and analysis, not on labor-intensive primer design and target selection. This kit can be easily automated on liquid handlers, further simplifying workflow.


Kit Specs

The NEXTflex Congenital Adrenal Hyperplasia Amplicon Panel contains enough material to prepare 8, 48, or 96 samples for Illumina® sequencing from DNA isolated from fresh or frozen samples. The shelf life of all reagents is 12 months when stored properly. All components can be safely stored at -20°C.

NEXTflex® Congenital Adrenal Hyperplasia Amplicon Panel

NEXTFLEX™ CAH Amplicon Panel Protocol

NEXTflex Congenital Adrenal Hyperplasia Amplicon Panel Kit Contents

NEXTflex® CAH Amplicon Primer Mix 1
NEXTflex® CAH Amplicon Primer Mix 2
NEXTflex® CAH Amplicon Primer Mix 3
NEXTflex® PCR Enhancer 1
NEXTflex® PCR Enhancer 4
NEXTflex® Hot Start PCR I Master Mix
NEXTflex® Ligation Mix
NEXTflex® Amplicon DNA Adapter
NEXTflex® PCR II Barcoded Primer Mix
NEXTflex® PCR II Master Mix
Nuclease-free Water
Resuspension Buffer
NEXTflex® Cleanup Beads

 

Required Materials Not Provided

20 - 100 ng of extracted genomic DNA (in up to 15.6 μL nuclease-free water each) 
Ethanol 80% (room temperature) 
96 well PCR Plate Non-skirted (Phenix Research™, Cat # MPS-499) or similar 
96 well Library Storage and Pooling Plate (Fisher Scientific®, Cat # AB-0765) or similar 
Adhesive PCR Plate Seal (Bio-Rad®, Cat # MSB1001) 
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 
Microcentrifuge 
1.5 mL nuclease-free microcentrifuge tubes 
Vortex

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For Use with DNA Isolated from FFPE Samples

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