|Volume for 500 ng||6.54||6.87||6.33||6.8||9.28||10.18||6.01||10.77||8.9||9.7||9.03||9.49|
|Water to Add||43.46||43.13||43.67||43.2||40.72||39.82||43.99||39.23||41.1||40.3||40.97||40.51|
|Final Volume (ul)||50||50||50||50||50||50||50||50||50||50||50||50|
Once elutided into 50 ul of nanopure water with 500 ng of DNA the samples were sonicated using the the sonication machine in the Seeb lab (make/model?) with the following protocol.
In 4C water
60 second on
30 seconds off
After that I waited until yesterday to do a quality check due to time constraints
I used the following protocol to check the gel.
Making a 1.3% agarose gel:
- 150 ml 1X TAE in a flask
- 2 g agarose mixed into TAE
- 5 ul SybrSafe mixed into Agarose
- Pour into mold with 15 well comb
- Allow gel to stiffen
Loading the Gel:
- 10 ul O’GeneRuler 100 bp Ladder in First and Last Well
- 20 ul sample mix with 5 ul 1X loading dye
- Load 25 ul dye/sample in each well
- 120 v for 2 hours
- Visualize with Dark reader with orange filter.
- Image for notebook
The samples were laid out on the gel in the following manner:
Ladder NF05 NF07 NF12 NF15 HL24 HL26 HL28 HL31 SN01 SN04 SN08 SN15 Ladder
After talking with Carita, Ryan, and Garret today I decided not to move forward with the bisulfite treatment and sequencing because of the poor quality of DNA.