Soft and Hard Homogenization Extractions

Soft and Hard Homogenization DNA/RNA Extractions #475-484

20190930 E.S. DNA/RNA Extractions from Montipora capitata and Pocillopora acuta adult coral fragments from Holobiont Integration Hawaii 2018 project.

Soft and Hard homogenization, and DNA/RNA Extractions followed this protocol. General Zymo Duet DNA/RNA Extraction protocol found here. With the following changes:

  • DNA: The first and second elutions were spun down in separate microcentrifuge tubes to compare the quantity and quality between the first and second elution. The first elution had no incubation period (previously 5 minutes) with 10 μl of Tris buffer and the second elution had a 15 minute incubation period (previously 5 minutes) with 100 μl of Tris buffer.
  • 10 μl aliquots of DNA and 5 μl aliquots of RNA (instead of 10 μl of each).

Montipora samples were atypically vortexed for 1 minute instead of 2 minutes during the soft homogenization step.

Qubit used to check DNA and RNA quantity (ng/μl).
TapeStation used to check RNA quality.

RNA Qubit Results: 20191001
DNA Qubit Results: 20191002

DNA Standard 1: 149.59 ng/μl
DNA Standard 2: 20,884.81 ng/μl
RNA Standard 1: 404.99 ng/μl
RNA Standard 2: 10,128.77 ng/μl

Timepoint Species Coral ID Extraction ID Homogenization DNA Reading 1 DNA Reading 2 Average DNA ng/μl RNA Reading 1 RNA Reading 2 Average RNA ng/μl RIN
20181215 Montipora 2380 475 soft 15.2 15.2 15.2 24.2 24.2 24.2 9
20181215 Montipora 2380 476 hard 7.54 7.5 7.52 19.4 19.4 19.4 **
20181215 Montipora 1178 477 soft 11.9 11.9 11.9 24.6 24.6 24.6 **
20181215 Montipora 1178 478 hard 14.6 14.5 14.55 18.8 18.8 18.8 9.2
20181215 Montipora 1076 479 soft 19 18.8 18.9 22.2 22.2 22.2 7.6
20181215 Montipora 1076 480 hard 13.5 13.3 13.4 17.2 17.2 17.2 **
20181215 Pocillopora 1777 481 soft 46.6 46.4 46.5 65.8 65.8 65.8 7.8
20181215 Pocillopora 1777 482 hard 30 29.8 29.9 32.4 32.6 32.5 7.3
20181215 Pocillopora 2879 483 soft 43.2 43 43.1 94 94.2 94.1 8.4
20181215 Pocillopora 2879 484 hard 35.8 35.6 35.7 57.4 57.6 57.5 8.1

Link to 20190930 TapeStation report, Extractions #475-484

Gel Electrophoresis Results:

20190930 Extractions #475-484

Written on September 30, 2019