Total RNA was extracted from tissues using TRIzol reagent (Invitrogen, catalog number: 15596026) according to the manufacturer’s instructions. Total RNA was processed to remove the genome DNA by using RQ1 RNase-Free DNase (Promega, catalog number: M6101). Then, 1 μg of RNA was reverse transcribed using M-MuLV Reverse Transcriptase Reaction system (NEB, catalog number: M0253L). cDNAs obtained were diluted and used for quantitative PCR (qPCR). Each qPCR assay was performed with a standard dilution curve of a calibrator, which was a mixture of different cDNA, to precisely quantify relative transcript levels. Gene-specific primers were used with SYBR green (Promega, catalog number: A6002) for detection on a LightCycler 480 system (Roche). The primer sequences used are synthesized by BGI as shown below:
M mulv reverse transcriptase reaction system
Manufactured by New England Biolabs
The M-MuLV Reverse Transcriptase Reaction system is a laboratory tool used for the conversion of RNA to complementary DNA (cDNA). It contains the M-MuLV reverse transcriptase enzyme, which catalyzes the synthesis of single-stranded cDNA from an RNA template.
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4 protocols using m mulv reverse transcriptase reaction system
1
Quantitative Analysis of Gene Expression
2
Quantitative RNA Expression Analysis
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Total RNA was extracted from the gastrocnemius, testis, caput epididymis and cauda epididymis using TRIzol reagent (Invitrogen, cat. no. 15596026) according to the manufacturer’s instructions. Total RNA was processed to remove genome DNA using RQ1 RNase-Free DNase (Promega, cat. no. M6101). Then, 1 μg of RNA was reverse transcribed using the M-MuLV Reverse Transcriptase Reaction system (NEB, cat. no. M0253L). cDNAs obtained were diluted and used for quantitative PCR (qPCR). Each qPCR assay was performed with a standard dilution curve of a calibrator, for a mixture of different cDNA, to precisely quantify relative transcript levels. Gene-specific primers were used with SYBR green (Promega, cat. no. A6002) for detection on a LightCycler 480 system (Roche). The primer sequences used were synthesized by Genewiz as shown in Supplementary Tab. 6 .
3
RNA Extraction and qRT-PCR Analysis
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Total RNA was extracted from hippocampus or liver using TRIzol reagent was performed as previously described [29 (link)]. One microgram of RNA was reverse transcribed using the M-MuLV Reverse Transcriptase Reaction system (NEB, cat. no. M0253L). Gene-specific primers were used with SYBR green (Promega, cat. no. A6002) for detection on a LightCycler 480 system (Roche). The primer sequences used were synthesized by BGI as shown Table S2 .
4
Quantitative Analysis of Gene Expression
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Total RNA was extracted from tissues using TRIzol reagent (Invitrogen, catalog number: 15596026) according to the manufacturer’s instructions. Total RNA was processed to remove the genome DNA by using RQ1 RNase-Free DNase (Promega, catalog number: M6101). Then, 1 μg of RNA was reverse transcribed using M-MuLV Reverse Transcriptase Reaction system (NEB, catalog number: M0253L). cDNAs obtained were diluted and used for quantitative PCR (qPCR). Each qPCR assay was performed with a standard dilution curve of a calibrator, which was a mixture of different cDNA, to precisely quantify relative transcript levels. Gene-specific primers were used with SYBR green (Promega, catalog number: A6002) for detection on a LightCycler 480 system (Roche). The primer sequences used are synthesized by BGI as shown below:
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