Description
General description
KAPA Mouse Genotyping Kits include KAPA Express Extract, a novel thermostable protease and buffer system that allows for the extraction of PCR-ready DNA from mouse tissue in as little as 15 minutes, and KAPA2G Fast Genotyping Mix with dye, containing a DNA polymerase engineered via directed evolution for high processivity and extreme speed. The combination of KAPA Express Extract and KAPA2G Fast Genotyping Mix allows for the reliable extraction and amplification of DNA fragments from mouse tissue in as little as 1 hour, as compared to ≥1 day with conventional protocols.KAPA2G Fast Genotyping Mix (2X) is a ready-to-use master mix containing all components for fast PCR, except primers and templates.
Application
KAPA Mouse Genotyping Kit has been used in the DNA extraction and amplification from mouse tail, ear, and toe tissue. The kit is also suitable for extraction and amplification of DNA from other animal tissue, but protocol optimization may be required.
Biochem/physiol Actions
DNA fragments generated with KAPA2G Fast Genotyping Mix are 3′-dA-tailed and may be cloned into TA cloning vectors, or used for routine downstream analyses or applications, including restriction enzyme digestion, cloning, and sequencing. Like wild-type Taq, KAPA2G Fast has 5′→3′ polymerase and 5′→3′ exonuclease activities, but no 3′→5′ exonuclease (proofreading) activity. The fidelity of KAPA2G Fast is similar to that of wild-type Taq; it has an error rate of approximately 1 error per 1.7 x 105 nucleotides incorporated.
Features and Benefits
Increased throughput, turnaround time and reliability:
PCR-ready DNA generated in as little as 15 minutes
Minimal handling, thereby reducing the risk of sample loss or contamination
Sufficient template for multiple assays; easily scaled to handle samples in a 96-well format
Fast and reliable amplification of DNA fragments across a wide range of amplicon lengths and GC contents
Outperforms crude extraction methods and commercial kits:
Increased turnaround times Increased specificity Decreased cost
Quick Notes:
Extract PCR-ready DNA from mouse tail, ear, or toe tissue in 15 minutes.
Dilute DNA extract 10-fold prior to PCR in 10 mM Tris-HCl (pH 8.0–8.5), and use 1 μL of DNA per 25 μL reaction.
Extracts are stable at -20°C for >6 months.
KAPA2G Fast (HotStart) Genotyping Mix contains the engineered KAPA2G Fast DNA Polymerase for fast PCR, along with buffer, MgCl₂ (1.5 mM at 1X), and dNTPs (0.2 mM each at 1X).
Loading dye for electrophoresis is included in the KAPA2G Fast Genotyping Mix, which allows direct loading of PCR products onto agarose gels for analysis.
Use 15 sec annealing time, with 15 sec extension time for amplicons
Preparation Note
Always ensure that the product has been fully thawed and mixed before use. Reagents may be stored at 4°C for short-term use (up to 1 month). Return to -20°C for long-term storage. Provided that all components have been handled carefully and not contaminated, the kit is not expected to be compromised if left (unintentionally) at room temperature for a short period of time (up to 3 days).
Analysis Note
All components contained with KAPA Mouse Genotyping Kits are subjected to stringent quality control tests, are free of contaminating exo- and endonuclease activities and meet strict requirements with respect to DNA contamination.
Other Notes
For Research Use Only. Not for use in diagnostic procedures.
