Table of Contents
What is m MLV Reverse Transcriptase?
Moloney Murine Leukemia Virus Reverse Transcriptase (M-MLV RT) is an RNA-dependent DNA polymerase that can be used in cDNA synthesis with long messenger RNA templates (>5kb). The enzyme is a product of the pol gene of M-MLV and consists of a single subunit with a molecular weight of 71kDa.
What is first strand synthesis?
Recommendations. Recommendations. The SuperScript™ First-Strand Synthesis System for RT-PCR is optimized to synthesize first-strand cDNA from purified poly(A)+ or total RNA. The system can be used with as little as 1 ng or as much as 5 µg of total RNA.
What is RT Mmlv?
M-MLV Reverse Transcriptase is a recombinant DNA polymerase that synthesizes a complementary DNA strand from single-stranded RNA, DNA, or an RNA:DNA hybrid. Compared to AMV RT, Moloney Murine Leukemia Virus Reverse Transcriptase (M-MLV RT) lacks DNA endonuclease activity and has a lower RNase H activity.
Which of the following genes is responsible for the coding of reverse transcriptase?
The pol gene encodes the three enzymes—protease, reverse transcriptase, and integrase—that catalyze the steps of retroviral infection.
How is cDNA different from a gene?
The main difference between DNA and cDNA is that DNA is composed of both coding and non-coding sequences whereas cDNA only contains the coding sequences. The coding sequences are the exons of a gene, which codes for a functional protein. The non-coding sequences are the remaining DNA sequences of the genome.
What is second strand cDNA synthesis?
Invitrogen Second Strand cDNA Synthesis Kit is designed to produce double-stranded cDNA from the first-strand reaction without the need for intermediate organic extraction or ethanol precipitation steps. The convenient single-tube format speeds up the synthesis procedure and maximizes cDNA recovery.
What are oligo dT primers?
Oligo dT primers are oligonucleotides that contain a segment of repeating deoxythymidines (dT). The dT anneal to the polyadenosine (polyA) tails of messenger RNA (mRNA), guiding the synthesis of complementary DNA (cDNA). Oligo dT primers are well-established reagents in cDNA synthesis and RT-PCR.
Why is Oligo dT an effective primer for reverse transcriptase?
Oligo(dT) primers amplify only mRNAs containing a poly(A) tail, since that is where the primer binds to promote reverse transcription. Random primers amplify most RNA species, including degraded RNA and viral genomes.
How do you find the cDNA of a gene?
- Finding cDNA sequence for a gene. Step 1 – Search. Step 2 – Choose a transcript. Step 3 – Access the cDNA sequence.
- Using a sequence to find a gene (BLAST/BLAT) Step 1 – Using BLAST/BLAT. Step 2 – View the results. Step 3 – Viewing the hit.
What is formed by reverse transcriptase?
A reverse transcriptase (RT) is an enzyme used to generate complementary DNA (cDNA) from an RNA template, a process termed reverse transcription.
What is Invitrogen M-MLV reverse transcriptase?
Invitrogen™ M-MLV Reverse Transcriptase. Recombinant DNA polymerase that synthesizes a complementary DNA strand from single-stranded RNA, DNA, or an RNA:DNA hybrid.
What is the recommended temperature for reverse transcription with M-MLV?
When using GeneAmp RNA PCR kit (part number N808-0017) with M-MLV, the recommended temperature for reverse transcription is 42°C. It is not recommended that reverse transcription be done at a higher temperature with this enzyme because it will start to lose significant activity.
What is one unit of M-MLV RT?
One unit of M-MLV RT is the amount of enzyme required to incorporate 1 nmole of deoxyribonucleotide into acid-precipitable material in 10 min at 37°C using poly (A)oligo (dT) 25 as template-primer.