Revision as of 21:05, 18 December 2014 by >Mzcai
CA12k_Nov2014 Probe Preparation
- Probe Design
- Received 190-mer oligos 12/18/14
- Suspended in 80ul TE Buffer
- Name: Fisga_Nov2014_seq
- Conc: 52.86 ng/ul (4.23 ug total)
- Calculate MW of 190 nt probe = 190nt * 308.9D/nt + 79D = 58,770 g/mol
- 52.86 ng/ul * pmol/58.770 ng = 0.8994 pmol/ul = ~0.9 uM
Aliquot
- Aliquot 40ul into 1.5ml tube
- From the aliquot take 1ul and dilute with H2O to 22.5ul in 1.5ml tube (40nM final conc)
900nM x 1ul = 40nM x 22.5ul
Expansion PCR Test
Component
|
Volume
|
Seed oligo |
0
|
F/R V6 Primer Mix (10uM) |
2
|
2X Kapa MM |
12.5
|
H2O |
10.5
|
Total |
25
|
Component
|
Volume
|
V6 +ve (2nM) |
10.5
|
F/R V6 Primer Mix (10uM) |
2
|
2X Kapa MM |
12.5
|
H2O |
0
|
Total |
25
|
Component
|
Volume
|
Seed oligo (40nM) |
0.6
|
F/R V6 Primer Mix (10uM) |
2
|
2X Kapa MM |
12.5
|
H2O |
9.9
|
Total |
25
|
Component
|
Volume
|
Seed oligo (40nM) |
6.25
|
F/R V6 Primer Mix (10uM) |
2
|
2X Kapa MM |
12.5
|
H2O |
4.25
|
Total |
25
|
Component
|
Volume
|
Seed oligo (900nM) |
2.8
|
F/R V6 Primer Mix (10uM) |
2
|
2X Kapa MM |
12.5
|
H2O |
7.7
|
Total |
25
|