qPCR with SYBR® Green .
Our new primaQUANT CYBR ADVANCED qPCR Master Mix with SYBR® Green was developed from scratch and shines in all essential performance criteria. Its high sensitivity (detection limit 1 template), wide dynamic range and high robustness guarantee reproducible results, even under difficult conditions.
The primaQUANT CYBR ADVANCED master mixes are very temperature stable. They do not need to be pipetted on ice and can easily be stored at room temperature for several days or in the refrigerator for several weeks.
The primaQUANT CYBR ADVANCED Master Mixes replace the primaQUANT CYBR Master Mixes with part numbers SL-9902, SL-9902R, SL-9902HR, SL-9902B, SL-9902RB, SL-9902HRB.
Hard Facts .
With the primaQUANT CYBR qPCR Master Mix, you can perform genotyping using High-Resolution Melting Curves (HRM) – without the need for agarose gels.
Li, W., Matsuoka, M., Kai, M., Thapa, P., Khadge, S., Hagge, D. A., Brennan, P. J., & Vissa, V. (2012). Real-Time PCR and High-Resolution Melt Analysis for Rapid Detection of Mycobacterium leprae Drug Resistance Mutations and Strain Types. Journal of Clinical Microbiology, 50(3), 742-753. https://doi.org/10.1128/JCM.05183-11
Słomka, M., Sobalska-Kwapis, M., Wachulec, M., Bartosz, G., & Strapagiel, D. (2017). High Resolution Melting (HRM) for High-Throughput Genotyping-Limitations and Caveats in Practical Case Studies. International Journal of Molecular Sciences, 18(11), 2316. https://doi.org/10.3390/ijms18112316
Lefever, S., Rihani, A., Van der Meulen, J., Pattyn, F., Van Maerken, T., Van Dorpe, J., Hellemans, J., & Vandesompele, J. (2019). Cost-effective and robust genotyping using double-mismatch allele-specific quantitative PCR. Scientific Reports, 9(1), 2150. https://doi.org/10.1038/s41598-019-38581-z
Single nucleotide polymorphism (SNP) genotyping is widely used in genetic studies to determine the factors underlying inherited traits.
Baris, I., Etlik, O., Koksal, V., Ocak, Z., & Baris, S. T. (2013). SYBR green dye-based probe-free SNP genotyping: introduction of T-Plex real-time PCR assay. Analytical Biochemistry, 441(2), 225-231. https://doi.org/10.1016/j.ab.2013.07.007
Dhas, D. B. B., Ashmi, A. H., Bhat, B. V., Parija, S. C., & Banupriya, N. (2015). Modified low cost SNP genotyping technique using cycle threshold (Ct) & melting temperature (Tm) values in allele specific real-time PCR. The Indian Journal of Medical Research, 142(5), 555-562. https://doi.org/10.4103/0971-5916.171282
Pang, Y., Zhou, Y., Wang, S., Lu, J., Lu, B., He, G., Wang, L., & Zhao, Y. (2011). A novel method based on high resolution melting (HRM) analysis for MIRU-VNTR genotyping of Mycobacterium tuberculosis. Journal of Microbiological Methods, 86(3), 291-297. https://doi.org/10.1016/j.mimet.2011.05.016
Bai, L., Deng, Y.-M., Dodds, A. J., Milliken, S., Moore, J., & Ma, D. D. F. (2006). A SYBR green-based real-time PCR method for detection of haemopoietic chimerism in allogeneic haemopoietic stem cell transplant recipients. European Journal of Haematology, 77(5), 425-431. https://doi.org/10.1111/j.1600-0609.2006.00729.x
Weksberg, R., Hughes, S., Moldovan, L., Bassett, A. S., Chow, E. W., & Squire, J. A. (2005). A method for accurate detection of genomic microdeletions using real-time quantitative PCR. BMC Genomics, 6, 180. https://doi.org/10.1186/1471-2164-6-180
The length of telomeres is well suited as a biomarker for cell aging. This also allows studying the impact of environmental influences on cell aging processes. qPCR is a rapid, reliable, and inexpensive method to study telomere lengths.
Lin, J., Smith, D. L., Esteves, K., & Drury, S. (2019). Telomere length measurement by qPCR – Summary of critical factors and recommendations for assay design. Psychoneuroendocrinology, 99, 271-278. https://doi.org/10.1016/j.psyneuen.2018.10.005
High Resultion Melting Curve Analysis (HRM) makes it easy to detect the smallest changes in the genome, such as SNPs and mutations, using qPCR.
Matsuda, K. (2017). PCR-Based Detection Methods for Single-Nucleotide Polymorphism or Mutation: Real-Time PCR and Its Substantial Contribution Toward Technological Refinement. Advances in Clinical Chemistry, 80, 45-72. https://doi.org/10.1016/bs.acc.2016.11.002
Simko, I. (2016). High-Resolution DNA Melting Analysis in Plant Research. Trends in Plant Science, 21(6), 528-537. https://doi.org/10.1016/j.tplants.2016.01.004
Mohammad Rahimi, H., Pourhosseingholi, M. A., Yadegar, A., Mirjalali, H., & Zali, M. R. (2019). High-resolution melt curve analysis: A real-time based multipurpose approach for diagnosis and epidemiological investigations of parasitic infections. Comparative Immunology, Microbiology and Infectious Diseases, 67, 101364. https://doi.org/10.1016/j.cimid.2019.101364
Next-generation sequencing requires accurate quantification of the DNA library to be sequenced for optimal results and high yields.
Such DNA libraries can be quantified inexpensively, rapidly and accurately by qPCR. By avoiding unnecessarily long PCR amplifications, the NGS workflow can be optimized, and amplification biases are also minimized.
Buehler, B., Hogrefe, H. H., Scott, G., Ravi, H., Pabón-Peña, C., O’Brien, S., Formosa, R., & Happe, S. (2010). Rapid quantification of DNA libraries for next-generation sequencing. Methods (San Diego, Calif.), 50(4), p15-18. https://doi.org/10.1016/j.ymeth.2010.01.004
Copy number variants can be measured by SYBR-Green qPCR using different Cq values after a calibration. In addition, the melting curve information allows an estimation of whether there are SNPs within the CNVs.
Loewe, R. P. (2013). Combinational usage of next generation sequencing and qPCR for the analysis of tumor samples. Methods (San Diego, Calif.), 59(1), 126-131. https://doi.org/10.1016/j.ymeth.2012.11.002
Diagnosis of pathogens can also be performed by nucleic acid analysis.
DNA pathogens (DNA viruses, bacteria, fungi) can be analyzed directly by quantitative PCR. For RNA pathogens (RNA viruses, e.g. SARS-COV2), a transcription of RNA into cDNA is necessary. For this purpose, we offer the primaREVERSE RT cDNA synthesis kit. Alternatively, you can perform cDNA synthesis and qPCR in one step with our primaQUANT 1-STEP Master Mixes.
Due to the particularly high purity of our primaQUANT Master Mixes, they can be used for quality control, e.g. to exclude the possibility that a product is contaminated with nucleic acids or nucleases.
Excellent performance .
The primaQUANT ADVANCED master mixes offer a wide dynamic range and excellent performance, even with low template amounts or targets with low expression.
Robust quantification even at low Cq values at the detection limit.
Wide Dynamic Range with excellent efficiency.
No template inhibition for large amounts of input