DNA/RNA extractions from E5 project

Extractions from the three coral species from each of the four timepoints

---

Extraction Date: October 08, 2021

Samples

Tube number	Timepoint	Species	Colony ID	Coll date	Site
23	January	Porites	POR-236	20200103	2
39	January	Acropora	ACR-244	20200103	2
73	January	Pocillopora	POC-200	20200103	2
421	March	Porites	POR-82	20200303	1
433	March	Acropora	ACR-343	20200304	3
457	March	Pocillopora	POC-378	20200304	3
523	Sept	Porites	POR-251	20200910	2
607	Sept	Porites	POR-383	20200908	3
615	Sept	Porites	POR-391	20200908	3
839	November	Porites	POR-341	20201031	3
847	November	Pocillopora	POC-377	20201031	3
853	November	Acropora	ACR-343	20201031	3

Tube 73 had a cap that said 73, but the side of the tube said 74

Extraction notes

• ACR and POC samples: pulled out 300ul of shield
• POR samples: pulled out 150ul of shield and added to 150ul of new shield
• Spun down samples for 3 minutes at 9000 rcf and then transfer the supernatant to new tube without disturbing the pellet
• 300ul of shield, 15ul of ProK, and 30ul of ProK digestion buffer, let sit for 2 minutes
• All spins were done for 1 minute or 2.30 minutes
• Did two washes with 700ul of wash buffer for both the DNA and RNA
• Then followed the protocol as described in protocol

Qubit

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

DNA

Tube number	RFU	DNA 1 (ng/uL)	DNA 2 (ng/uL)	Average
Standard 1	197.80
Standard 2	23245.31
23		nd	nd	nd
39		31.4	30.8	31.1
73		59.4	59.0	59.2
421		3.46	3.36	3.41
433		24.6	23.8	24.2
457		33.6	33.4	33.5
523		4.10	4.00	4.05
607		26.4	26.6	26.5
615		13.5	13.6	13.55
839		6.02	5.82	5.92
847		2.48	2.36	2.42
853		45.2	44.6	44.9

RNA

Tube number	RFU	RNA 1 (ng/uL)	RNA 2 (ng/uL)	Average
Standard 1	372.52
Standard 2	6731.91
23		13.4	13.2	13.3
39		14.0	13.4	13.7
73		42.6	42.4	42.5
421		23.6	24.2	23.9
433		14.4	13.6	14.0
457		29.6	30.4	30.0
523		14.6	14.8	14.7
607		21.2	21.0	21.1
615		37.6	37.6	37.6
839		15.0	14.8	14.9
847		49.0	48.8	48.9
853		25.0	24.8	24.9

Tape Station

• Used to check RNA quality Protocol
• Results Link

Gel

• Modified from this protocol
• Added 0.75g of agarose and 50ml of 1x TAE to flask and microwaved for 45 seconds. This makes a 1.5% gel
• Once cool enough to touch added 2ul of gel green stain
• Swirled and poured into gel mould with comb
• Once solidified, covered with 1X TAE as a running buffer
• Added 1ul of purple loading dye to each of my QC strip tube samples. I had ~9ul of DNA leftover from QC and ~8ul of RNA
• Loaded my gel with the DNA first, then skipped a well and then the RNA
• Ran the gel for 60 minutes at 60 volts

Addtional Notes

• NA