For Research Use Only.

Figure: Amplification of human GAPDH gene target with 2X HoTaq Real-time PCR Kit. Amplification curves are shown for ten fold dilutions of 0.0002pM to 20pM of plasmid. Inset shows the standard curve data.

Description:
This is a high performance real-time PCR reagent. It utilizes MCLAB's proprietary quantitative PCR technology.

Advantages:
2x HoTaq PCR Reaction Mix products are superior in amplifying difficult templates comparing with similar products from other suppliers.

- This is the amplification of GPIIB gene (70% G+C) .
- 10 ~ 10K copies from 30pg human genomic DNA have been detected.

Applications:
Probe based quantitative PCR: including DNA quantification, 2-step RT PCR, SNP analysis, etc.

Primer and probe design:
1. To achieve the best performance, appropriate software, such as ABI Primer Express^TM, should be used to design primers with 50°C~65°C annealing temperature and 68°C~70°C for probes with 17~30 nucleotides in length
2. Amplicon size should be small, <150bp
3. Avoid secondary structures in primers and probes
4. Avoid more than 3 consecutive Gs in primers and probes
5. Primers should not have complementary 3' -ends
6. 17 ~ 30 nucleotides in length

Recommended Reaction Conditions: :
95°C, 10 minutes. -> (95°C, 5 seconds. -> 60°C, 30 seconds.) for 50 cycles.

Recommended Storage Condition:-20°C

Notes:
To achieve accurate quantification, it is highly recommended to do replicates and to reduce pipetting errors.

Reference:
1. Holland, P. M., Abramson, R. D., Watson, R., and Gelfand,
2. D. H. 1991. Proceedings of the National Academy of Sciences USA 88:7276-7280.
3. Livak, K. J., Flood, S. J. A., Marmaro, J., Giusti, W., and Deetz, K. 1995. PCR Methods and Applications 4:357-362.
4. Lee, L. G., Connell, C. R., and Bloch, W. 1993 Nucleic AcidsResearch 21:3761-3766.