component
STS-MRT10
STS-MRT50
M-MLV Reverse T.
10000 units / 50ul
50000 units / 250ul
5X Reverse Buffer
1ml 5X Buffer
1ml 5X Buffer
DTT
500ul / 100mM
500ul / 100mM
First Strand cDNA synthesis
Assay Set-Up:
Before starting, vortex all components thoroughly to ensure homogeneity.
Prepare a premix for the number of assays you need according to the following protocol:
First - Strand cDNA synthesis
Final volume: 20ul
- In a nuclease-free microcentrifuge tube add:
- dNTPs mix (10mM each) 1ul
- 1ng to 4ug total RNA
or
1-500ng mRNA xul
- 50-100ng random primers
or
Oligo(dT)12-18 (500ug/ml) 1ul
- ddH2O up to 12ul
- Heat mixture @ 65°C for 2min. and quick cool the tube on ice. Add:
- 5X Reverse Buffer 4ul
- 100mM DTT 2ul
- ddH2O 1ul
or
Ribonuclease Inhibitor
(optional)
- Mix gently and incubate
- @ 42°C for 2 min. if you are using oligo(dT)12-18
or
- @ 25°C for 2 min. if you are using random primers
- Add 1ul (200U) of GeneSpin M-MLV Reverse Transcriptase
(if you are using random primer incubate @25°C for 10 min.)
- Incubate @ 42°C for 60 - 120 min.
- Inactivate the reaction @ 70°C for 10 min.
GeneSpin Srl
Via Friuli, 51
20135 Milano
P.IVA C.F. 04520270960
Laboratory
and sales office
c/o Fondazione Filarete
Viale Ortles, 22/4
20139 Milano
tel. +39 02 56660199
fax. +39 02 537250
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.
cat. no.
amount
note
STS-MRT10
10000 units
5X Reverse Buffer
STS-MRT50
50000 units
5X Reverse Buffer
STS-MRT200
200000 units
5X Reverse Buffer
Reverse Transcriptase
5X Master Mix
Primers
Oligonucleotides
FOR RESEARCH USE ONLY
UNIT DEFINITION
One unit is defined as the amount of enzyme required to incorporate 10 nanomoles of dNTPs into acid-insolubile material in 30 min at 74°C.
SHIPPING
Shipped in green ice.
STORAGE
Store at -20C°
SHELF LIFE
12 months
FORM
Liquid
CONCENTRATION
200U/ul
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administration@pec.genespin.com