---
author: Sam White
toc-title: Contents
toc-depth: 5
toc-location: left
date: 2015-07-27 23:23:24+00:00
layout: post
slug: dnase-treatment-o-lurida-ctenidia-1hr-post-mechanical-stress-rna
title: DNase Treatment - O.lurida Ctenidia 1hr Post-Mechanical Stress RNA
categories:
  - 2015
  - Olympia oyster reciprocal transplant
tags:
  - ctenidia
  - DNase
  - gill
  - NanoDrop1000
  - olympia oyster
  - Ostrea lurida
  - RNA
  - RNA quantification
  - Turbo DNA-free
---

Quantified the RNA I isolated from Jake's samples on [20150715](https://robertslab.github.io/sams-notebook/posts/2015/2015-07-16-rna-isolation-o-lurida-ctenidia-1hr-post-mechanical-stress-2//robertslab.github.io/sams-notebook/2015/07/10/rna-isolation-o-lurida-ctenidia-1hr-post-mechanical-stress/) using the Roberts Lab NanoDrop1000 (ThermoFisher).

![](https://eagle.fish.washington.edu/Arabidopsis/20150727_Oly_ctenidia_RNA_mech_stress_ODs.JPG)(http://eagle.fish.washington.edu/Arabidopsis/20150727_Oly_ctenidia_RNA_mech_stress_ODs.JPG)



![](https://eagle.fish.washington.edu/Arabidopsis/20150727_Oly_ctenidia_RNA_mech_stress_plots_01.JPG)(http://eagle.fish.washington.edu/Arabidopsis/20150727_Oly_ctenidia_RNA_mech_stress_plots_01.JPG)



![](https://eagle.fish.washington.edu/Arabidopsis/20150727_Oly_ctenidia_RNA_mech_stress_plots_02.JPG)(http://eagle.fish.washington.edu/Arabidopsis/20150727_Oly_ctenidia_RNA_mech_stress_plots_02.JPG)



Overall, the yields are good. The 260/280 ratios are mediocre. Will proceed with DNase treatment.

DNased 1.5ug of RNA from each sample using the Turbo DNA-free Kit (Ambion/Life Technologies), following the “rigorous” protocol.

Briefly:




    
  * 50μL reactions were carried out in 0.5mL tubes

    
  * added 1μL of DNase to each tube

    
  * incubated 30mins @ 37C

    
  * added additional 1μL of DNased

    
  * incubated 30mins @ 37C

    
  * added 0.2 vols (10.2μL) of DNase Inactivation Reagent

    
  * incubated and mixed for 2mins @ RT

    
  * spun 1.5mins, 10,000g @ RT

    
  * transferred 50μL of supe to sterile 1.5mL snap cap tubes

    
  * spec’d on Roberts Lab NanoDrop1000



Samples were stored @ -80C in [Shellfish RNA Box #6](https://docs.google.com/spreadsheets/d/1ax6C-muxUTXxFEtfWdswBvueLhmxZzmwZcO2ur-0q-Q/edit?usp=sharing). Will quantify at a later date.

DNase reaction calcs: [20150727_Jake_Oly_mech_stress_DNase_calcs](https://docs.google.com/spreadsheets/d/1o5Ri2POi4R-ZsPGQvT4AFgl6CK8lsdID74GwE7ZJjFI/edit?usp=sharing)




