FAST PGT-A PREP: 3 HOURS FROM CELL LYSIS TO NGS READY
PG-Seq™ Rapid Kit v2 for Streamlined PGT-A Preparation
for Illumina® Instruments
The PG-Seq™ Rapid kit v2 has been developed to analyze picogram quantities of DNA (single/multi-cells or low template DNA) from an embryo biopsy for preimplantation genetic testing. The kit utilizes whole genome amplification (WGA) and next generation sequencing (NGS) technology to accurately screen all 24 chromosomes for whole chromosome aneuploidy and sub-chromosomal abnormalities. From DNA to data, the kit includes all reagents required for cell lysis, whole genome amplification, indexing along with the PG-Find™ analysis software for automatic calling of aneuploidy and copy number variants.
4340-0248
48 RXNS
4341-IL48A
ILLUMINA® BARCODES 1-48 | 48 RXNS
4341-IL48A
ILLUMINA® BARCODES 49-96 | 48 RXNS
UPDATED STREAM-LINED, SIMPLE PROTOCOL
Accurate Copy Number Detection
Extensively tested using over 100 cell line and genomic DNA samples with known ploidy.
- Tested from whole chromosome aneuploidies down to segmental aberrations 7Mb in size with 30pg of genomic DNA or 5 cell fibroblast samples, representative of a trophectoderm biopsy.
- See PG-Seq™ Rapid kit v2 app note for more information.
Flexible Kit Format
- 48rxn format with two sets of 48 unique indexes available, allowing up to 96 sample multiplexing (up to 384 sample multiplexing available by custom request).
- Larger 40 µL WGA PCR 1 reaction volume to enable options for alternative downstream processing.
- High WGA PCR 1 yield of 2-4ug total DNA with only 23 cycles of PCR.
Improved Whole Genome Coverage & Accuracy
- Enhanced PG-Find quality scores compared to the v1 kit which signifies less noise, less bias and higher confidence (Figure 1).
- Improved whole genome coverage compared to the v1 kit with a 70% increase in the number of targets covered when PG-Seq™ Rapid v2 WGA PCR 1 product is used in hybridization capture panels.
Figure 1: Comparison of quality scores obtained from cell line samples processed through PG-Seq™ Rapid v1 and v2. A lower quality score indicates less noise and more reliable results.
Figure 2: IGV coverage track of the full mitochondrial genome (chrM: 1-16364bp) from a PG-Seq™ Rapid v2 amplified 5-cell sample with 1,000,000 sequencing reads.
High % Coverage of the mtDNA Genome
- > 90% coverage of the mtDNA genome with no protocol modifications (Figure 2).
- Ability to analyze and track embryo SNPs for sample tracing purposes.
Easy-to-Use, Highly Customizable Analysis Software
- Two analysis options, self-reference and reference based, to accommodate every laboratory’s individual needs.
- Multiple visualization features including selecting which chromosomes to view and report, a single check box to view the raw vs smoothed data, unlimited color options, multiple result images.
- User adjustable event calling thresholds that can be used to designate mosaic, 1 copy or 2 copy gains and losses
- An easy to interpret result overview screen allows the results of every sample to be easily analyzed and for general data trends to be observed.
- A modifiable binning distance allowing the CNV resolution to be tailored according to laboratory requirements and sample sequencing depth and coverage.
- Automatic and manual copy number calling of whole chromosome and sub-chromosome copy number events, including detailed information on the position and size of each event.
Kit Contents
- Cell Lysis Mix
- PCR 1 Ready-mix
- PCR 1 Primer
- PCR 2 Ready-mix
- Clean-up Beads
- Resuspension Buffer
INDEXED PRIMERS
- Indexed primers, suitable for Illumina® Sequencing instruments
Pre-PCR Laboratory requirements
- Illumina® MiSeq® or MiniSeq® sequencer
- Illumina® sequencer related consumables
- Laminar flow cabinet
- Minicentrifuge
- Pipettes (P2, P10, P20, P100, P200, P1000 µl)
- Cold block (4°C)
- Thermocycler (with hotlid & programmable ramp rate to 0.3°C/sec)
- Pipette tips (low binding, barrier filter)
- PCR thin walled reaction tube with flat cap (0.5 mL or 0.2 mL)
- Sterile, molecular grade tubes (1.5 mL)
Post-PCR Laboratory requirements
- Agarose gel-electrophoresis apparatus
- Electrophoresis power supply
- UV transilluminator or gel documentation instrument
- Multi-channel pipette
- Multi-channel pipette reagent reservoirs
Illumina® sequencer reagent kit (select from):
- Illumina® MiSeq® Reagent Kit v3 (150 cycle, cat # MS-102-3001)
- Illumina® MiSeq® Reagent Kit v2 (300 cycle, cat # MS-102-2002)
- Illumina® MiSeq® Reagent Micro Kit v2 (300 cycle, cat # MS-103-1002)
- Illumina® MiSeq® Reagent Nano Kit v2 (300 cycle, cat # MS-103-1001)
- Illumina® MiniSeq® High Output Reagent Kit (75 cycles, cat # FC-420-1001)
NGS SUPPORT INFORMATION
PG-Seq™ Rapid Kit for Illumina® NGS
- Sequences of PG-Seq™ Rapid Kit Indexes – Excel / PDF
- Illumina® Samplesheet
MANUAL
TECHNICAL NOTE
The PG-Seq™ Rapid kit v2 contains enough reagents to prepare 48 samples for Illumina® Sequencers. The shelf life of all reagents is at least 6 months when stored according to the specified storage temperatures.
Avila Perez, C., Parnell, L., Florensa Bargallo, M., Herreros Cuesta, J., Larreategui Laiseca, Z., Prados Dodd, N., Ruiz Perez, M., & Wells, D. (2023). P-731: The accuracy of truly non-invasive PGT using spent culture media is insufficient to justify routine clinical use. Human Reproduction, 38(Supplement_1), dead093. 1050.
Banu, M., Pathan, A. A. K., & Chaitanya, K. V. (2023). Diagnostics for Genetically Inherited Disorders: From Cytogenetics to Genomics Technologies-A Review. Biomedical and Pharmacology Journal, 16(2).
Cheng, H. Y. H., Chow, J. F. C., Lam, K. K. W., Lai, S. F., Yeung, W. S. B., & Ng, E. H. Y. (2023). Randomised double-blind controlled trial of non-invasive preimplantation genetic testing for aneuploidy in in vitro fertilisation: a protocol paper. BMJ Open, 13(7), e072557.
Curnow, E., Ryan, G. L., & Yu, B. (2023). Discordant non-invasive PGT-A results and clinical outcome. Fertility and Sterility, 120(4), e276.
Spath, K., Costa-Borges, N., Nikitos, E., Kostaras, K., Calderón, G. C., Psathas, P., & Wells, D. (2023). P-730: Detection of mitochondrial reversal following meiotic spindle transfer: a finding of importance for mitochondrial replacement therapies used for the purpose of avoiding…. Human Reproduction, 38 (Supplement_1), dead093. 1049.
Sonehara, H., Matsumoto, R., Nakayama, N., Kobanawa, M., Numata, K., Kawasaki, A., & Shozu, M. (2022). Aneuploidy and sex concordance rate between cell-free DNA analysis from spent culture media of preimplantation embryo and DNA from whole embryo with respect to different…. Reproductive Medicine and Biology, 21(1), e12493.