---
author: Sam White
toc-title: Contents
toc-depth: 5
toc-location: left
layout: post
title: DNA Shearing & Bioanalyzer - Lotterhos C.virginica Mantle gDNA from 2018114
date: '2019-03-06 15:26'
tags:
  - gDNA
  - shearing
  - Crassostrea virginica
  - mantle
  - Eastern oyster
categories:
  - 2019
  - Miscellaneous
---
_Crassostrea virginica_ mantle gDNA that had previously been [isolated on 20181114](https://robertslab.github.io/sams-notebook/posts/2018/2018-11-14-DNA-Isolation-and-Quantification-Lotterhos-C.virginica-Mantle-DNA/) was evaluated for integrity via agaroase gel [yesterday (20190305)](https://robertslab.github.io/sams-notebook/2019/03/05/Agarose-Gel-Lotterhos-C.virginica-Mantle-DNA-from-20181114/). Everything looked pretty good, so proceeded with shearing in preparation for MBD enrichment.

Aliquoted 2500ng of DNA in a final volume of 100uL of each sample in 0.5mL snap cap (polypropylene) tubes for shearing in the Seeb Lab sonicator (Diagenode Bioruptor 300) with the following settings:

- 30 cycles

- 30 seconds on

- 30 seconds off

![Cycle settings on Diagenode Bioruptor for DNA shearing](https://github.com/RobertsLab/sams-notebook/blob/master/images/20190306_bioruptor_settings_DNA_shearing.jpg?raw=true)

Calculations for DNA dilutions are here (Google Sheet):

- [Roberts_2017AdultExposureSampleInfo](https://docs.google.com/spreadsheets/d/1cDLmp6jKf37gnPTwHDR07dNt-t3_jzGE2TC2afagouM/edit?usp=sharing)

Ideally, this will shear DNA to a target size around 300bp.

After shearing, ran all samples on the Bionalyzer 2100 (Agilent) using a DNA 12000 chip according to manufacturer's protocol.


---

# RESULTS

Date files (XAD - requires Windows and 2100 Expert software):

- [2100 expert_DNA 2012000_DE72902486_2019-03-06_09-33-02.xad](http://owl.fish.washington.edu/Athaliana/20190306_virginica_bioanlyzer/2100%20expert_DNA%2012000_DE72902486_2019-03-06_09-33-02.xad)

- [2100 expert_DNA 12000_DE72902486_2019-03-06_10-12-36.xad](http://owl.fish.washington.edu/Athaliana/20190306_virginica_bioanlyzer/2100%20expert_DNA%2012000_DE72902486_2019-03-06_10-12-36.xad)

Electropherograms:

![Bioanalyzer DNA12000 electopherograms of sheard DNA 1 of 2](http://owl.fish.washington.edu/Athaliana/20190306_virginica_bioanlyzer/20190306_virginica_bioanalyzer_electropherogram_all-01.jpg)

![Bioanalyzer DNA12000 electopherograms of sheard DNA 2 of 2](http://owl.fish.washington.edu/Athaliana/20190306_virginica_bioanlyzer/20190306_virginica_bioanalyzer_electropherogram_all-02.jpg)


Overall, shearing was decent, albeit, the majority of the resulting fragementation is a tad bit on the larger side (e.g. ~500bp). However, all of these samples will be subjected to bisulfite conversion which should fragment the DNA further, so things should be fine.
