Quantitative real-time PCR (qRT-PCR) assay

WH Weiguo He
HL Huiqing Liu
LH Linlin Hu
YW Yaohui Wang
LH Lane Huang
AL Aihong Liang
XW Xuan Wang
QZ Qing Zhang
YC Yi Chen
YC Yi Cao
SL Suyun Li
JW Junli Wang
XL Xiaocan Lei
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Total RNA was extracted from the testes of rats with TRIzol reagent (Themo Fisher, Inc., USA). cDNA was synthesized with the TransScript II One-Step gDNA Removal and cDNA Synthesis SuperMix kit (Transgen Biotech, Beijing, China) according to the manufacturer’s protocol. By using GAPDH as the internal reference, quantitative PCR analyses were carried out in the SYBR Green assay system with the Applied Biosystems 7500 Real-time PCR System (Applied Biosystems, Foster City, CA, USA). The mRNA expression levels were calculated by using the 2-ΔΔCt method. The primers used in this study were listed in Table 1.

Primer pairs and corresponding annealing temperatures used in this study

F: GCTCCATCCTGAAGAAGGT

R: TGCACTAAGGAGACGTGAGA

F:GTGGCAGTAACAGTGACAGTG

R:GTCAGCTCCAGATCCTCCAG

F:GATGGAATGGAGCAGAAGAC

R: CACAATCGTAACTGGTCAGC

F: ACAACTCCCAGAAGCCTAAGAATG

R: RGCTTTTCCCTTGGCAGCTATG

F: GGAGAATGGCAAGAATGAAGA

R: RCCGCAGGAAGGTAAAGAG

F: CGAGCATGGGTTCCATGTC

R: CTGGACCGCCATGTTTCTTAG

F: CCTCAAGATTGTCAGCAATG

R: CAGTCTTCTGAGTGGCAGTG

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