---
author: Sam White
toc-title: Contents
toc-depth: 5
toc-location: left
layout: post
title: Reverse Transcription - C.gigas RNA from Noah's Heat-Mechanical Stress Project
date: '2023-07-13 12:48'
tags: 
  - reverse transcription
  - Crassostrea gigas
  - RNA
  - Pacific oyster
categories: 
  - 2023
  - Miscellaneous
---
After [isolating ctenidia RNA on 20230712](https://robertslab.github.io/sams-notebook/posts/2023/2023-07-12-RNA-Isolation-C.gigas-Ctenidia-from-Noah's-Heat-Mechanical-Stress-Project/) and then quantifying the RNA [earlier today](https://robertslab.github.io/sams-notebook/2023/07/13/RNA-Quantification-C.gigas-RNA-from-Noah's-Heat-Mechanical-Stress-Project/), I needed to perform reverse transcription to have cDNA for subsequent qPCRs. Reverse transcription was performed using oligo dT primers using M-MLV RT (Promega), per the manufacturer's recommendations. Used 100ng of RNA in each reaction. All reactions were done on ice in 0.5uL PCR tubes.

- [20230713-cgig-ctenidia-Noah-cDNA-calcs](https://docs.google.com/spreadsheets/d/1tVS6JLsSJlU-NCJvhhyeHfpW4eTchfSgX-mlOEnHda4/edit?usp=sharing) (Google Sheet)

In preparation for qPCR and primer testing, 2uL from each sample was pooled together in a separate 0.5uL tube. All cDNA was stored in Noah's -80<sup>o</sup>C project box.