Applied Biosystems 7300 Getting Started Guide

Chapter 3 Performing Reverse Transcription
Converting Total RNA to cDNA
Applied Biosystems 7300/7500/7500 Fast Real-Time PCR System Relative Quantification Getting Started Guide 19
Notes
Primer Extended on mRNA
5′
3′
Reverse
Primer
5′
Synthesis of 1st cDNA strand
cDNA
Oligo d(T) or random hexamer
3′
5′
cDNA
3
Converting Total RNA to cDNA
Using the High
Capacity cDNA
Archive Kit
Use the High Capacity cDNA Archive Kit (PN 4322171) to perform the first step (RT)
in the two-step RT-PCR method. Follow the manual method for converting total RNA
into cDNA, as specified in the High Capacity cDNA Archive Kit Protocol (PN 4322169).
IMPORTANT! The protocol is not shipped with the High Capacity cDNA Archive Kit.
Download the protocol from
http://docs.appliedbiosystems.com/search.taf
To search for the document, select ABI P
RISM
®
6100 Nucleic Acid PrepStation in the
Product list box, then click Search at the bottom of the page. The protocol is listed under
the Protocols heading.
Thermal Profile
Parameters for RT
The High Capacity cDNA Archive Kit uses the following thermal profile parameters for
the RT step.
Note: Thermal cycling conditions for one-step RT-PCR are described on page 30.
Step Type Time Temperature
HOLD 10 min 25 ° C
HOLD 120 min 37 ° C

Applied Biosystems 7300 Specifications

General

Brand	Applied Biosystems
Model	7300
Category	Laboratory Equipment
Language	English