Acropora Larvae DNA/RNA Extraction Batch 3

Larvae from FACE PUF Morea Project


For detailed extraction protocol view post: Protocol for DNA/RNA Extractions of Montipora Coral Larvae Using Zymo-Duet Extraction Kit


Extraction Date: August 20, 2019

Samples

Extraction Number Sample ID Content Sample Quantity (µL) Sample Conserv. Tech
21 014 Sperm 100 300 uL Shield Erin
22 755 Sperm 100 300 uL Shield Erin

Extraction Notes

  • Altough samples were already preserved in 300µL DNA/RNA Sheild, I added an additional 300µL of shield to prevent RNA/DNA degradation during digest
  • After adding 300µL shield I transferred the samples to a 1.7mL centrifuge tube in order to digest for 1.5 hours, 55°C, 1100 rpm
  • Added additional 15µL Proteinase K after 2 hours
  • To accomodate the use of extra shield, we added 750µL Lysis buffer instead of 345µL (Step 3 DNA Extraction). Additionally, we mixed 1000µL EtOH instead of 700µL in 5µL tubes (Step 1 RNA Extraction) and repeated steps 6-7 once.

Qubit

  • Followed Broad Range dsDNA and RNA Qubit protocol
  • All samples were read twice
Extraction # Sample ID DNA Standard 1 (RFU) DNA Standard 2 (RFU) DNA 1 (ng/µl) DNA 2 (ng/µl) Average DNA RNA Standard 1 (RFU) RNA Standard 2 (RFU) RNA 1 (ng/µl) RNA 2 (ng/ul) Average RNA
21 14 164.0 18126 60.8 60.6 60.7 390.6 9983 Too Low Too Low Too Low
22 755 164.0 18126 47.0 46.8 46.9 390.6 9983 34.4 34.4 34.4

Tape Station

  • Followed RNA TapeStation protocol
  • Only Tape Stationed 755

TS-Morea-FACE-PUF-Batch-3-Ext-Ladder

TS-Morea-FACE-PUF-Batch-3-Ext-22

Gel

  • Followed PPP Lab Agarose Gel protocol for a medium 1% agarose gel
  • Used the Thermo Scientific GeneRuler 1 kb Plus DNA Ladder (link)

annotated-Morea-FACE-PUF-gel-batch-3.png

Notebook Entry 20190820
Lab_Notebook_20190820_Morea-FACE-PUF-batch-3

Written on August 20, 2019