Soft and Hard Homogenization Extractions

20191113 M.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).

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

High Sensitivity:
DNA Standard 1: 41.06 ng/μl
DNA Standard 2: 20,514.38 ng/μl
RNA Standard 1: 51.27 ng/μl
RNA Standard 2: 1,003.4 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
20180922 Pocillopora 2534 559 soft 69.6 69.2 69.4 39.4 39.6 39.5 6.9
20180922 Pocillopora 2534 560 hard 38.6 38.6 38.6 19.6 19.6 19.6 NA
20180922 Montipora 1609 561 soft ** ** ** 34 34 34 8.2
20180922 Montipora 1609 562 hard 42.4 42.4 42.4 ** ** ** NA
20180922 Montipora 1628 563 soft 48 47.8 47.9 44.8 45.2 45 8.1
20180922 Montipora 1628 564 hard 26.2 26.2 26.2 22.2 22.2 22.2 NA
20180922 Pocillopora 2733 565 soft 48.6 48.6 48.6 23.6 23.6 23.6 6.7
20180922 Pocillopora 2733 566 hard 32.8 32.8 32.8 14.1 14.2 14.15 NA
20180922 Montipora 1169 567 soft 40.8 40.8 40.8 46.2 46.2 46.2 7.9
20180922 Montipora 1169 568 hard 28.4 28.4 28.4 26.4 26.4 26.4 NA
20180922 Montipora 1289 569 soft 24 23.8 23.9 14.1 14.1 14.1 8.6
20180922 Montipora 1289 570 hard 14.3 14.3 14.3 7.92 7.9 7.91 NA
20181117 Montipora 1204 571 soft 14.3 14.3 14.3 5.46 5.42 5.44 **
20181117 Montipora 1204 572 hard 9.68 9.66 9.67 5.4 5.42 5.41 NA
20180922 Montipora 1124 573 soft 27.8 27.8 27.8 16.7 16.6 16.65 8.8
20180922 Montipora 1124 574 hard 18.9 18.9 18.9 10.2 10.2 10.2 NA

Link to 20191115 TapeStation report, Extractions #539-558

Gel Electrophoresis Results:

20191115 Extractions #558-574

Written on November 13, 2019