RNA and DNA Extractions for ENCORE Project, April 18, 2023

Protocol Link

Extraction of 18 samples from ENCORE Project. These samples were collected in Bermuda in August 2022 for the ENCORE TPC project. There are 160 samples total from the following 3 species: Diploria labyrinthiformis (DLAB), Montastraea cavernosa (MCAV), and Madracis decactis (MDEC) from the following temperatures 16, 22, 26, 29, 33, 36, and 40 ºC

Samples

Continuing on with the rest of the ENCORE extractions.

Notes

• Followed protocol exactly. Took 300 uL from the sample tubes for all species before proceeding with the extraction as written.
• The DLAB DNA and RNA concentrations have been consistenly low but the quality is good. Eluted all DLAB DNA in RNA in 50 uL instead of 100 uL.

Qubit Results

• Used Broad range dsDNA and RNA Qubit Protocol
• All samples read twice, standard only read once

DNA Standards: 191.24 (S1) & 21938.4 (S2)

colony_id	Species	Temp.Cat	DNA_QBIT_1	DNA_QBIT_2	DNA_QBIT_AVG
Dlab-A7	Diploria labyrinthiformis	36	8.08	8.46	8.27
Dlab-B6	Diploria labyrinthiformis	26	4.72	4.94	4.83
Dlab-B7	Diploria labyrinthiformis	36	9.52	9.78	9.65
Dlab-C5	Diploria labyrinthiformis	22	3.66	3.8	3.73
Dlab-D2	Diploria labyrinthiformis	40	25.2	25.8	25.5
Dlab-D7	Diploria labyrinthiformis	36	30.2	30.8	30.5
Dlab-E2	Diploria labyrinthiformis	40	18.8	19.1	18.95
Dlab-F7	Diploria labyrinthiformis	36	36.6	37.2	36.9
Mcav-A6	Montastraea cavernosa	26	6.76	6.8	6.78
Mcav-B2	Montastraea cavernosa	40	86.6	87.6	87.1
Mcav-B7	Montastraea cavernosa	36	43	44.6	43.8
Mcav-C5	Montastraea cavernosa	22	5.22	5.34	5.28
Mcav-C6	Montastraea cavernosa	26	4.96	4.9	4.93
Mcav-D3	Montastraea cavernosa	33	5.86	5.98	5.92
Mdec-B6	Madracis decactis	26	6.22	6.36	6.29
Mdec-B7	Madracis decactis	36	16.5	16.7	16.6
Mdec-D1	Madracis decactis	29	8.4	8.38	8.39
Mdec-F5	Madracis decactis	22	5.9	5.98	5.94

RNA Standards: 422.04 (S1) & 8600.79 (S2)

colony_id	Species	Temp.Cat	RNA_QBIT_1	RNA_QBIT_2	RNA_QBIT_AVG
Dlab-A7	Diploria labyrinthiformis	36	10.6	10.4	10.5
Dlab-B6	Diploria labyrinthiformis	26	16	14.8	15.4
Dlab-B7	Diploria labyrinthiformis	36	13.4	14	13.7
Dlab-C5	Diploria labyrinthiformis	22	13.4	13.4	13.4
Dlab-D2	Diploria labyrinthiformis	40	21.2	21.2	21.2
Dlab-D7	Diploria labyrinthiformis	36	19.4	19.8	19.6
Dlab-E2	Diploria labyrinthiformis	40	25.2	25	25.1
Dlab-F7	Diploria labyrinthiformis	36	17.8	18.2	18
Mcav-A6	Montastraea cavernosa	26	15.4	14.8	15.1
Mcav-B2	Montastraea cavernosa	40	26.8	27.4	27.1
Mcav-B7	Montastraea cavernosa	36	24	22.6	23.3
Mcav-C5	Montastraea cavernosa	22	10.4	10	10.2
Mcav-C6	Montastraea cavernosa	26	0	0	0
Mcav-D3	Montastraea cavernosa	33	19.6	19.2	19.4
Mdec-B6	Madracis decactis	26	18.8	19	18.9
Mdec-B7	Madracis decactis	36	14.8	15	14.9
Mdec-D1	Madracis decactis	29	26.6	28	27.3
Mdec-F5	Madracis decactis	22	15.2	15	15.1

DNA and RNA Quality Check: Gel, using Kristina’s gel protocol at the bottom of this page.

Next steps

Will continue on with the rest of the extractions for these 3 species at these temps! 34 + any re-extractions to go! Hoping to finish by the end of the semester.

Still holding off on the Porites astreoides extractions until we can find a suitable protocol.