qPCR with SYBR® Green .

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

To determine the expression of a gene (gene or RNA expression), the total RNA must first be transcribed into cDNA.

For separate cDNA synthesis, our primaREVERSE RT Kit is the first choice.

Alternatively, you can also use our primaQUANT 1STEP One-Step RT-qPCR Kit and directly use RNA in a qPCR.

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

More information:

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.