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MMLV Reverse Transcriptase

货号

M6125H

规格

2,500 U @ 10 U/μl

存储温度

-20℃

产品详情

Applications

First-strand cDNA synthesis for subsequent PCR or real-time PCR.
RT-PCR validation of gene expression data obtained from microarray experiments.
RT-PCR validation and quantification of gene silencing by RNA interference.

The MMLV High Performance Reverse Transcriptase (MMLV HP RT) and the MMLV Reverse Transcriptase 1st-Strand cDNA Synthesis Kit both produce full-length first-strand cDNA from total cellular RNA preparations or purified poly(A) RNA.

MMLV HP RT demonstrates significantly greater reverse transcriptase activity than other commercially available MMLV RT enzymes. Typically, just 100 units of MMLV HP RT are required for full-length cDNA synthesis compared to 200 units of competitive MMLV RT enzymes. MMLV HP RT includes a 10X Reaction Buffer, optimized for synthesis of full-length cDNA from long RNA templates, and DTT.

Benefits

Synthesize full-length cDNA from RNA templates longer than 15 kb.
MMLV HP RT demonstrates significantly higher activity than competitive reverse transcriptase enzymes.
Reaction Buffer, optimized for producing full-length cDNA, is included with both the MMLV HP RT and the MMLV Reverse Transcriptase 1st-Strand cDNA Synthesis Kit.
The kit includes both an oligo(dT) and random nonamer primers.
First-strand cDNA can be made from picogram amounts of input total RNA.
The kit includes a potent RNase Inhibitor to protect the integrity of template RNA.

Figure 1. MMLV HP RT produces full-length cDNA from mRNA longer than 15 kb. Total RNA isolated from HeLa cells was reverse transcribed and the cDNA was amplified by PCR. Detection of the 1.3-kb PCR amplicon from near the 5´ end of the mRNA demonstrates full-length reverse transcription of HERC1 mRNA (A). Agarose gel analysis of the PCR products shows the 1.3-kb amplicon from the 5´ end of the mRNA (B). Lane M, 100 bp DNA ladder; lane 1, no-RT control reaction; lane 2, PCR product from cDNA synthesized by EPICENTRE's MMLV HP RT.

Target Transcript (Size)	3´/5´ Ratios
Target Transcript (Size)	EPICENTRE MMLV High Performance Reverse Transcriptase	Competitor I (RNase H-Mutant of MMLV RT)	Competitor P (MMLV RT)
ACTB (1,792 b)	0.9	1.7	1.2
GUSB (2,162 b)	1.0	6.1	2.5
TFRC (5,010 b)	5.5	12.1	11.3

Table 1. 3´/5´ ratio analysis of cDNA produced by different reverse transcriptase enzymes. Total cellular RNA from HeLa cells was converted to cDNA using the three reverse transcriptase enzymes indicated in the table. A 3´/5´ ratio equal to 1.0 means that equal amounts of PCR products are obtained from both the 3´ and 5´ end of the cDNA and therefore is a good indication that the reverse transcriptase has produced a full-length cDNA copy of the mRNA.

2A.

Figure 2. cDNA produced by EPICENTRE's MMLV High Performance Reverse Transcriptase yields a significantly improved 3´/5´ ratio than competitive reverse transcriptases. The approximately 2160-base HeLa β-glucuronidase mRNA (GUSB) was reverse transcribed into cDNA using EPICENTRE's MMLV High Perfomance Reverse Transcriptase and two competitive reverse transcriptase enzymes. PCR primer pairs to the 3´-end and 5´-end of GUSB cDNA were synthesized and qPCR (SYBR^® Green I dye detection) was performed using each primer pair and the GUSB cDNAs as templates. The 3´/5´ ratio was calculated for each as described in the text. (A). PCR amplicons from the 3´ end and 5´ end of the GUSB cDNA. (B). qPCR quantification graphs for detecting the 3´ amplicon and 5´ amplicon of GUSB cDNA produced by EPICENTRE's MMLV High Performance Reverse Transcriptase Kit and two other commercially available reverse transcriptase enzymes.

	5'-amplicon
	3'-amplicon

2B.

EPICENTRE MMLV 1st-Strand cDNA Synthesis Kit

Company I RNase H- minus MMLV RT

Company P MMLV RT

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