---
author: Sam White
toc-title: Contents
toc-depth: 5
toc-location: left
layout: post
title: Agarose Gel - Lotterhos C.virginica Mantle DNA from 20181114
date: '2019-03-05 13:27'
tags:
  - Crassostrea virginica
  - eastern oyster
  - mantle
  - gDNA
  - gel
categories:
  - 2019
  - Miscellaneous
---
Checked DNA integrity of the _Crassostrea virginica_ mantle gDNA I [isolated on 20181114](https://robertslab.github.io/sams-notebook/posts/2018/2018-11-14-DNA-Isolation-and-Quantification---Lotterhos-C.virginica-Mantle-DNA/) in preparation for MBD enrichment. Detailed sample info and sample processing (including calculations for MBD enrichment using the MethylMiner Kit) are here (Google Sheet):

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

Loaded 100ng of each sample on a 0.8% agarose, 1X Low TAE gel with ethidium bromide. Each sample was mixed with 3uL of 6x loading dye and brought up to 18uL with nuclease-free H<sub>2</sub>O. Gel was run for ~2hrs @ 100V. Used two ladders in a (failed) attempt to accurately assess gDNA size:

- [GeneRuler HighRange DNA Ladder](https://github.com/RobertsLab/resources/blob/master/protocols/Commercial_Protocols/ThermoFisher_GeneRuler_HighRange_DNALadder_50ug.pdf) (PDF)

- [GeneRuler DNA Ladder Mix](https://github.com/RobertsLab/resources/blob/master/protocols/Commercial_Protocols/ThermoFisher_OgeneRuler_DNA_Ladder_Mix_F100439.jpg) (JPG)

The idea being that the HighRange ladder would handle high integrity gDNA molecular weight, while the DNA Ladder Mix would handle any degraded DNA and/or RNA present.

---

# RESULTS

Wish I had wiped the dust off of the protective UV screen before snapping the pic...

![20190305_gel_gDNA_virginica](https://github.com/RobertsLab/sams-notebook/blob/master/images/20190305_gel_gDNA_virginica.jpg?raw=true)

Almost all of the samples look pretty good. The exceptions are:

- 17013
- 17176
- 17203

These three samples appear to be degraded (evidenced by lack of prominent, high molecular weight band).

Also, just to add info regarding the ladders, the HighRange ladder's smallest band is ~10kbp and the largest is ~48kbp.

The DNA Ladder Mix's smallest band is ~100bp and the largest is ~10kbp.

The DNA Ladder Mix ran from the upper lane into the lane below it, giving the appearance of a continuous ladder; however, it is just overlapping the DNA Ladder Mix in the lane below. Additionally, the HighRange ladder did not have sufficient time to separate out in to defined bands - probably needed a couple of more hours!

Will proceed with shearing tomorrow morning and then MBD enrichment.


![GeneRuler HighRange DNA Ladder](https://github.com/RobertsLab/resources/blob/master/protocols/Commercial_Protocols/ThermoFisher_GeneRuler_HighRange_DNALadder_50ug.png?raw=true)

![GeneRuler DNA Ladder Mix](https://github.com/RobertsLab/resources/blob/master/protocols/Commercial_Protocols/ThermoFisher_OgeneRuler_DNA_Ladder_Mix_F100439.jpg?raw=true)