Relative abundance Pocillopora spp. around the island of Mo’orea

Context:

Our study is based on population genetics of Pocillopora sp. on the Polynesian islands based on their often misleading and incorrect morphology. We are therefore trying to understand population dynamics by Haplotype so that we can correlate our transect data with our term tolerance data from the laboratory (see NoteBook TPC Haplotype).

gDNA extraction with “Quick-DNA 96 Kit / Cat No: D3012

Protocol:

1. Reagent preparation: Add 260ul or 1,040ul Proteinase K storage Buffer to reconstitute the lyophilized Proteinase K, 5mg (D3001-2-5) or 20mg (D3001-2-20), respectively (final concentration of 20mg/ml). Vortex to dissolve.

2. gDNA extraction in DNA/RNA shield

   • Add 10ul PK Digestion buffer + 5ul of Proteinase K.
     For 2 plate in a time –> 2.0ml of PK Digestion Buffer + 1.0ml ProtK

   • Add 400ul og Genomic Lysis Buffer to 100ul of teh sample/shield mixture.
     For 2 plate in a time –> 400ul x 192 samples = 76.8ml

   • Mix completely by vortexing (with pipette in the plate) 4-5 times then let stand 10 minutes

   • Transfer the mixture to the wells of a Silicon-A Plate on a Collection Plate. Centrifuge at 2.500xg for 5 minutes.

   • Remove the waste in the colelction plate and move to the next step.

   • Add 200ul of DNA Pre-Wash Buffer to each well and. centrifuge at 2,500xg for 5 nminutes,
     For 2 plate in a time –> 200ul x 192 samples = 38.4ml.

   • Remove the waste in the colelction plate and move to the next step.

   • Add 300ul g-DNA Wash Buffer to each well and centrifuge at 2,500xg for 5 minutes.
     For 2 plate in a time –> 300ul x 192 samples = 57.6ml.

   • Transfer the Silicon-A Plate onto an Elution plate. Add 50ul DNA Elution Buffer (need to be heat at 70˚C) to each well and incubate during 2 minutes then centrifuge at 2,500xg for 5 minutes. For 2 plate in a time –> 50ul x 192 samples = 9.6ml.

   • Keep the Silicon-A Plate in the fridge (4˚C).

   • Cover the Elution Plate and keep it at -20˚C.

   • Nano drop 3-4 samples on each plate to control de DNA concentration.

Result:

20240919 Plate 16 and 17.

See plate number for each samples here.

Plate	Sample	Well	ng/ul	A260/280	A260/230
17	1398	A1	17	1.72	0.85
17	1390	B1	1.1	1.6	0.16
16	1407	H12	1.5	2.75	0.56
16	2220	A1	.- 0,8	0.84	.-0.06
16	2194	B1	.-0.9	1.16	.-0.25

After checking we figure it out that we didin’t add Elution Buffer but g-DNA Wash buffer. Following that we tooked plates out of the freezer and added 50ul of Elution Buffer Eluted in 50ul of Elution Buffer and mesured on nanodrop.

Plate	Sample	Well	ng/ul	A260/280	A260/230
16	2220	A1	3.4	1.97	0.89
16	2194	B1	20.8	1.88	1.83
16	2189	C1	13.6	1.9	1.96
17	1398	A1	10	1.82	1.01
17	1390	B1	14.9	1.86	2.32
17	2163	C1	12.1	1.91	1.31

20240920 Plate 18 and 19.

See plate number for each samples here.

Plate	Sample	Well	ng/ul	A260/280	A260/230
Elution buffer	x	x	0.1	.-0.14	0.18
18	2418	A1	15.6	2.11	2.3
18	2350	B1	23.8	2.05	0.33
18	2421	C1	31.2	2.03	1.64
19	1448	A1	16.9	2.1	2.16
19	1442	B1	12	2.14	2.22
19	1418	C1	4.1	2.83	0.56

20240920 Plate 20 and 21.

See plate number for each samples here.

Plate	Sample	Well	ng/ul	A260/280	A260/230
Elution buffer	x	x	0.1	.-1.64	0.58
20	1424	A1	14.3	2.08	2.05
20	2463	B1	33.2	1.97	1.46
20	3314	C1	28.1	1.94	1.58
21	2137	A1	5.6	2.11	1.04
21	2152	B1	10.4	1.97	2.22
21	3113	C1	24.4	1.97	2.02

Probability of mixing buble surface between 12C and 12D on plate 20.

20240920 Plate 22 and 23.

See plate number for each samples here.

Plate	Sample	Well	ng/ul	A260/280	A260/230
Elution buffer	x	x	0.8	.-12.88	2.26
22	2136	A1	18.1	1.94	1.48
22	3231	B1	39	1.95	1.89
22	3291	C1	22.8	1.98	1.64
23	3496	A1	25.8	2	2.46
23	3620	B1	19	1.98	1.65
23	2916	C1	37.6	1.93	2.42

20240921 Plate 24 and 25.

See plate number for each samples here.

Plate	Sample	Well	ng/ul	A260/280	A260/230
Elution buffer	x	x	0	.-0.01	0.12
24	3513	A12	61.8	1.96	2.33
24	3501	B12	34	2.02	1.75
24	3674	C12	30.8	2.01	1.76
25	2752	F12	21.8	2.05	2.45
25	2781	G12	20.1	2.07	2.26
25	2768	H12	43.4	2.01	2.27

20240922 Plate 26 and 27.

See plate number for each samples here.

Plate	Sample	Well	ng/ul	A260/280	A260/230
Elution buffer	x	x	0.2	2.78	0.43
26	2715	A1	39	1.93	1.72
26	2708	B1	35.4	1.9	0.93
26	2845	C1	32	1.95	1.55
27	3445	A12	42.9	1.92	0.84
27	3679	B12	8.9	2.11	1.83
27	2694	C12	15.9	1.98	1.4

20240922 Plate 28 and 29.

See plate number for each samples here.

Plate	Sample	Well	ng/ul	A260/280	A260/230
Elution buffer	x	x	.-0.4	1.3	.-5.44
28	2386	F1	33.5	1.91	1.94
28	2382	G1	38	1.93	1.9
28	2361	H1	35.9	1.92	1.88
29	2996	A1	34.1	1.95	1.6
29	2855	B1	42.1	1.9	1.09
29	2605	C1	35.2	1.95	2.14

20240922 Plate 30 and 31.

See plate number for each samples here.

Plate	Sample	Well	ng/ul	A260/280	A260/230
Elution buffer	x	x	.-0.1	0.27	.-0.95
31	3385	A12	31.9	1.93	2.05
31	3364	B12	29.1	1.94	2.02
31	3431	C12	10.5	1.83	0.53
30	2665	F12	35.8	1.91	2.18
30	2991	G12	28.5	1.94	1.63
30	2500	H12	47.9	1.9	1.81

20240923 Plate 32 and 33.

See plate number for each samples here.

Plate	Sample	Well	ng/ul	A260/280	A260/230
Elution buffer	x	x	.-0.1	0.18	.-0.18
32	430	A1	35.6	1.96	2.1
32	2085	B1	12.6	2.1	1.62
32	1975	C1	36.8	1.99	1.87
33	2004	A12	34.4	1.95	1.38
33	1914	B12	29.6	2	1.71
33	1779	C12	19.9	2.04	2.08

20240923 Plate 34 and 35.

See plate number for each samples here.

Plate	Sample	Well	ng/ul	A260/280	A260/230
Elution buffer	x	x	0.2	.-0.45	0.88
34	1530	A1	19.7	1.97	1.68
34	1659	B1	24.7	2.02	1.87
34	1567	C1	22.6	2.02	2.07
35	1527	F12	46.5	1.92	2.11
35	1928	G12	31.1	1.94	1.63
35	1985	H12	38.9	1.95	2.19

20240924 Plate 36 and 37.

See plate number for each samples here.

Plate	Sample	Well	ng/ul	A260/280	A260/230
Elution buffer	x	x	0.1	.-0.09	.-0.32
36	1959	A1	62.1	1.93	2.37
36	1963	B1	16.5	2.03	2.28
36	2025	C1	23.5	1.99	2.64
37	931	A10	54.4	1.96	2.13
37	1218	A11	32.7	1.93	2.7
37	1075	A12	47.8	1.93	1.81

20240925 Plate 38 and 39.

See plate number for each samples here.

Plate	Sample	Well	ng/ul	A260/280	A260/230
Elution buffer	x	x	0.3	.-0.30	.-1.36
38	814	A1	31	1.99	2.13
38	892	A2	45.1	1.97	1.8
38	799	A3	24.4	2.04	1.16