Pole Cancer Kit
The BioVendor fastGEN POLE Cancer kit prepares a sequencing library for the POLE gene by next-generation sequencing on an Illumina instrument. One kit covers 16 reactions, each with its own index. POLE encodes the catalytic subunit of DNA polymerase epsilon. Pathogenic variants in the exonuclease domain cause an ultramutated tumour phenotype, which is one of the four molecular groups in the classification of endometrial carcinoma and also occurs in colorectal cancer.
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This kit in detail
- POLE exons 9, 11, 13 and 14, eleven codons
- NGS library preparation in a single-step PCR, amplicons up to 170 bp
- 16 reactions per kit, each with its own index
- DNA from tumour tissue, including FFPE
- About 4 hours in total, under 30 minutes hands-on
- Illumina MiSeq or MiniSeq, analysis in GENOVESA
The BioVendor fastGEN POLE Cancer kit prepares sequencing libraries for genotyping the POLE gene by massively parallel sequencing. It is a library preparation kit, not a DNA extraction kit and not a real-time PCR assay: the DNA has to be isolated beforehand, and the result comes from the sequencer.
Regions covered
POLE, the catalytic subunit of DNA polymerase epsilon, in exons 9, 11, 13 and 14. The codons covered are 286, 295, 297, 367, 368, 411, 424, 436, 444, 456 and 459. The manufacturer catalogue number is RDNGS0004.
Method
The kit uses a single-step PCR with hybrid primers that produce short amplicons of up to 170 bp. Sample and Master Mix are combined in about 10 minutes, the reaction runs on a real-time PCR instrument, and the pooled and purified libraries are sequenced on an Illumina instrument. Total workflow time is around 4 hours, of which less than 30 minutes is hands-on.
Sample and DNA input
The input is 5 µl of isolated DNA per reaction, at 10 to 200 ng/µl measured by spectrophotometer or from 1 ng/µl measured with the Qubit HS assay. DNA from tumour tissue, including formalin-fixed paraffin-embedded material, is used.
Kit contents
The kit contains 16 ready-to-use Master Mixes, each with its own index, plus the read 1 sequencing primer (R1SP), the read 2 sequencing primer (R2SP) and the index sequencing primer (ISP). The sequencing primers are added to positions 12, 13 and 14 of the Illumina cartridge alongside the Illumina primers. No positive or negative control is supplied. Storage is at -20 °C.
Analysis
Raw FASTQ data is analysed in the fastGEN module of the GENOVESA software, a cloud application supplied for this purpose.
Regulatory status
The product data sheet for this kit states that it is for research use only.
- Master Mix, 16 vials, each with a unique index
- R1SP, read 1 sequencing primer
- R2SP, read 2 sequencing primer
- ISP, index sequencing primer
16 reactions per kit, REF RDNGS0004. No positive or negative control is supplied; the manufacturer recommends running controls of your own. DNA isolation reagents are not supplied. Storage at -20 °C until the expiry date. The sequencing primers are added to positions 12, 13 and 14 of the Illumina cartridge together with the Illumina primers.
Datasheets and regulatory documentation
Everything your quality team needs for evaluation and procurement.
Brochure
Application Protocol
Frequently paired with this kit
Other assays and instruments lab teams add alongside the Pole Cancer Kit.
Add this kit to your quote
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