---
author: Sam White
toc-title: Contents
toc-depth: 5
toc-location: left
date: 2015-10-30 00:48:23+00:00
layout: post
slug: dna-isolation-geoduck-olympia-oyster-2
title: DNA Isolation – Geoduck & Olympia Oyster
categories:
  - 2015
  - Geoduck Genome Sequencing
  - Olympia Oyster Genome Sequencing
tags:
  - DNA Isolation
  - DNA Quantification
  - gDNA
  - geoduck
  - NanoDrop1000
  - olympia oyster
  - Ostrea lurida
  - Panopea generosa
---

Amazingly, we need more gDNA for the two genome sequencing projects (geoduck and Olympia oyster). Used geoduck adductor muscle sample from Box 1 of the geoduck [samples collected by Brent & Steven on 20150811.](https://onsnetwork.org/halfshell/2015/08/11/big-day-big-clam/) Used Olympia oyster ctenidia from Box 1 of [adductor muscle sample collected by Brent & Steven on 20150812.](http://onsnetwork.org/halfshell/2015/08/12/another-day-another-species/)

Tissues were split in approximately half, minced and transferred to tubes with 1mL of DNAzol + 50μg/mL of Proteinase K (Fermentas). Previously, I had just homogenized samples. I'm hoping that the overnight digestion with Proteinase K will help increase yields from these.

Tissue weights:







    
  * Geoduck adductor muscle tube 1: 292mg (gone)

    
  * Geoduck adductor muscle tube 2: 320mg (gone)

    
  * Olympia oyster ctenidia tube 1: 135mg (gone)

    
  * Olympia oyster ctenidia tube 2: 130mg (gone)


Samples were isolated using DNAzol (Molecular Research Center) according to the manufacturer’s protocol, with the following adjustments:



    
  * Samples were incubated O/N @ RT on a rotator.

    
  * After Proteinase K digestion, added 40μL RNAse A (100mg/mL) and incubated @ RT for 15mins.

    
  * Performed optional centrifugation step (10,000g, 10mins @ RT)

    
  * Initial pellet wash was performed using a 70%/30% DNAzol/EtOH

    
  * Pellets were resuspended Buffer EB (Qiagen)


Resuspension volume = 500μL total for each species

Samples were incubated O/N at RT to facilitate pellet resuspension.

NOTE: Geoduck "pellets" were not very DNA pellet-like. Very loose, white, and sort of disintegrate (but not dissolve in solution) when attempted to resuspend.


