PCR 1 amplifying ITS2 marker for Box 1 - O.Fav deep

Protocol

Notes:

• Samples that were amplified with the previous protocol were re run with new protocol (see ITS2_PCR1_12)
• Samples were re diluted from 30uL aliquot stock DNA

Samples:

• OFAV = Orbicella faveolata
• OFRA = Orbicella franksi
• S = Shallow
• P = Deep

Species	Treatment	ID
OFAV	P	9GT46
OFAV	P	352
OFAV	P	356
OFAV	P	360
OFAV	P	369
OFAV	P	OHS3
OFAV	P	9AZ5
OFAV	P	DW

Master Mix:

Component	Volume for 1 rxn	Volume for 8 rxns (+1% error)
2x MyTaq Mix	25.0 uL	202 uL
Molecular Water	18.0 uL	145.4 uL
ITS2 Primer Forward (10 uM)	2.0 uL	16.2 uL
ITS2 Primer Reverse (10 uM)	2.0 uL	16.2 uL
DNA	3.0 uL	NA

Strip Tube Set Up:

Strip (replicate) Number	A	B	C	D	E	F	G	H
1	9GT46	352	356	360	369	OHS3	9AZ5	DW
1	9GT46	352	356	360	369	OHS3	9AZ5	DW
1	9GT46	352	356	360	369	OHS3	9AZ5	DW

Gel Map:

Failed: 9GT46, 352, 356, OHS3, 9SZ5

The next step is to use “cleaned DNA” Matias made for tricky samples