DNA Extraction of Oly Broodstock

• SS1_1
  SS1_2 (not tan)
  SS1_3
  SS1_4
  SS1_5
  SS1_6
  SS1_7
  SS1_8
  SS1_9
  SS1_10
  SS1_11
  SS1_12
  SS1_13
  SS1_14
  SS1_15
  SS1_16
  SS1_17
  SS1_18
  SS1_19
  SS1_20
  SS3_1
  SS3_2
  SS3_5
  SS3_6
• Found 2 tubes labelled SS1_2. Marked the unextracted one with a tan sticker
• Could not initially find SS3_3 or SS3_4 but found later and reorganized box
• Left in 60degC water bath for 3 hours
• Eluted 50 uL then 100 uL

Primers

• Received the rest of the 2b-RAD primers and resuspended them to 100 uM.
  • BC11
  • BC12
  • HT3
  • HT4
  • HT5
  • HT6
  • HT7
  • HT8
• Made new 1 uM aliquots of BC2-10 and HT1
  • 1 uL stock + 99 uL NFW
• Made new 10 uM  aliquots of Lib 1 and Lib 2
  • 2.5 uL stock + 22.5 uL NFW

PCR for 2b-RAD MiSeq run

Used 10 samples from 9/28/15 digestion and ligation. Numbers in parentheses are so I know which tube from the ligation each sample is in.

1. SS2_6 (5)
2. SS2_7 (6)
3. SS2_8 (7)
4. HC1_5 (8)
5. HC1_6 (9)
6. HC1_7 (10)
7. HC1_8 (11)
8. NF1_4 (14)
9. NF1_5 (15)
10. NF1_6 (16)

Master Mix

This time, set up a medium gel for 8 of the samples and a small gel for the other 2. Made a 2% TAE gel, and after running it at 110V for 1 hour let the gels soak in water with 6 uL of EtBr to increase staining. PCR used 19 cycles.

Gel of 2b-RAD PCR product

I think I should make sure to mix in the EtBr when doing the post staining as that is likely why there are bright white sections on the gels. It was still much easier to see the bands though. Cut out the bands and left them in 1.5 mL tubes in 4degC for gel extraction on Friday.