DNA/RNA extractions from E5 project

Re-Extractions from Acropora from the fourth timepoint plus one from TP one

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Extraction Date: December 22, 2022

Samples

Tube number	Timepoint	Species	Colony ID	Coll date	Site
864	November	Acropora	ACR-267	20201030	2
866	November	Acropora	ACR-244	20201030	2
872	November	Acropora	ACR-228	20201030	2
896	November	Acropora	ACR-234	20201030	2
912	November	Acropora	ACR-225	20201030	2
920	November	Acropora	ACR-265	20201030	2
930	November	Acropora	ACR-237	20201030	2
932	November	Acropora	ACR-229	20201030	2
936	November	Acropora	ACR-256	20201030	2
35	January	Acropora	ACR-223	20200103	2

Extraction notes

• For my samples I took 300ul the RNA/DNA shield and added it to a new 1.5ml tube containing 15ul of ProK, and 30ul of ProK digestion buffer
• Samples sat for 15 minutes at room temperate
• Spun down samples for 3 minutes at 9000 rcf and then transfer the supernatant to new tube without disturbing the pellet
• All subsequent 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	210.67
Standard 2	22923.60
864		40.2	39.0	39.6
866		73.2	70.4	71.8
872		31.0	30.4	30.7
896		52.8	51.6	53.6
912		70.2	70.0	70.1
920		78.2	76.8	78.8
930		204	204	204
932		84.4	83.4	83.9
936		94.6	93.2	93.9
35		27.4	26.6	27.0

RNA

Tube number	RFU	RNA 1 (ng/uL)	RNA 2 (ng/uL)	Average
Standard 1	426.92
Standard 2	9405.32
864		96.2	95.4	95.8
866		95.0	94.6	94.8
872		139	137	138
896		104	104	104
912		85.8	84.8	85.3
920		148	147	147.5
930		112	110	111
932		66.4	66.4	66.4
936		76.4	75.6	76.0
35		104	104	104

Tape Station

• Samples not analyzed

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

• The skeleton weirdly dissolved in the RNA/DNA shield. The sample turned clear after the centrifugation
• Everything still looks like garbage :(