Apr 15, 2025
Version 2
cDNA synthesis V.2
- Núria Peñuelas1,
- Miguel Chillon2,
- Miquel Vila1
- 1Vall d'Hebron Research Institute;
- 2Department of Biochemistry and Molecular Biology, Universitat Autònoma de Barcelona, 08193 Bellaterra, Spain. Institute of Neurosciences, Universitat Autònoma de Barcelona, 08193 Bellaterra, Spain. UAB-VHIR Joint Unit, Vall d’Hebron Institut de Recerca, 08035 Barcelona, Spain. Catalan Institution for Research and Advanced Studies (ICREA), 08010 Barcelona, Spain
- Vilalab Public
Protocol Citation: Núria Peñuelas, Miguel Chillon, Miquel Vila 2025. cDNA synthesis . protocols.io https://dx.doi.org/10.17504/protocols.io.kqdg3948qg25/v2Version created by Miquel Vila
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: September 13, 2022
Last Modified: April 15, 2025
Protocol Integer ID: 126724
Funders Acknowledgements:
ASAP
Grant ID: ASAP-020505
Abstract
High-Capacity cDNA Reverse Transcription Kits(#4368814, Applied Biosystems - ThermoFisher)
cDNA synthesis
cDNA synthesis
Prepare the 2X master mix (ul per sample):
- 2ul RT Buffer
- 0.8ul 25X dNTP Mix(100mM)
- 2ul Random primers
- 1ul Multiscribe Reverse Transcriptase
- 4.2ul Nuclease free water
Take 0.5ug of RNA in a maximum of 10ul of sample volume
Mix the 2X master mix (10ul) with the RNA samples (10ul) and mix by pippeting
Centrifuge the plates to spin down all the reagents.
Keep the samples on ice
Program the thermal cycler to 25C for 10min 37C for 120 mintues 85C for 5 minutes and 4C until finished.
Store the cDNA samples at -20C for short-term storage or -80C for long-term storage (years).