Noi/NOTES/2012-5-20: Difference between revisions
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'''Reagent prep''' | '''Reagent prep''' | ||
Add the following reagents to CT Conversion Reagent | Add the following reagents to CT Conversion Reagent | ||
- H2O 9.0mL | |||
- M-Dissolving Buffer 500ul | |||
- M-Dilution Buffer 3.0mL | |||
Mix at room temperature with frequent vortexing or shaking for 15 minutes | |||
Add 144 ml of 100% ethanol to the 36 ml M-Wash Buffer concentrate before use | |||
{| {{table}} border = 1 | {| {{table}} border = 1 | ||
Line 247: | Line 250: | ||
| Ma15-21||172.9||6.94||Ma15-21||H5||13.06||1200.00||130.00||150.00 | | Ma15-21||172.9||6.94||Ma15-21||H5||13.06||1200.00||130.00||150.00 | ||
|} | |} | ||
'''Program''' | |||
98°C for 10 minutes | |||
64°C for 2.5 hours | |||
4°C storage up to 20 hours | |||
Measure conc. with ssDNA Qubit assay (used 1ul of bis-cvt gDNA) |
Revision as of 17:33, 20 May 2012
DMR 330k BSPP capture of samples from Sergio
Sample list
New gDNA extracted from the cell pellet
on: http://genome-tech.ucsd.edu/LabNotes/index.php/Noi/NOTES/2012-5-17
No. | Sample ID | Lab ID | Conc. (ng/ul) | Volume (ul) | Yields (ug) |
1 | HUES2 p41 | Ap15-1 | 110.0 | 250 | 27.5 |
2 | ASThiPS4F1 p12 | Ap15-2 | 139.1 | 250 | 34.8 |
3 | ASThiPS4F2 p12 | Ap15-3 | 145.1 | 250 | 36.3 |
4 | HUES6 p39 Control TeSR | Ap15-4 | 295.5 | 250 | 73.9 |
5 | HUES6 p39 ACTIVIN | Ap15-5 | 128.1 | 250 | 32.0 |
6 | HUES6 p39 BMP-4 | Ap15-6 | 153.9 | 250 | 38.5 |
7 | HUES9 p39 Control TeSR | Ap15-7 | 183.5 | 250 | 45.9 |
8 | HUES9 p39 ACTIVIN | Ap15-8 | 149.3 | 250 | 37.3 |
9 | HUES9 p39 BMP-4 | Ap15-9 | 111.2 | 250 | 27.8 |
10 | HUES3 p36 Control TeSR | Ap15-10 | 153.5 | 250 | 38.4 |
11 | HUES3 p36 ACTIVIN | Ap15-11 | 87.5 | 250 | 21.9 |
12 | HUES3 p36 BMP-4 | Ap15-12 | 109.7 | 250 | 27.4 |
13 | H1 p54 Control TeSR | Ap15-13 | 191.3 | 250 | 47.8 |
14 | H1 p54 ACTIVIN | Ap15-14 | 108.7 | 250 | 27.2 |
15 | ASThiPS4F5 p16 Control TeSR | Ap15-15 | 134.2 | 250 | 33.6 |
*16 | ASThiPS4F5 p16 ACTIVIN | Ap15-16 | 50.2 | 500 | 25.1 |
17 | ASThiPS4F4 p36 Control TeSR | Ap15-17 | 183.0 | 250 | 45.8 |
18 | ASThiPS4F4 p36 ACTIVIN | Ap15-18 | 82.6 | 250 | 20.7 |
19 | ASThiPS4F4 p36 BMP-4 | Ap15-19 | 263.4 | 250 | 65.9 |
20 | H1 p48 | Ma15-2 | 215.2 | 200 | 43.0 |
21 | H9 p47 | Ma15-3 | 252.0 | 200 | 50.4 |
22 | HUES3 p33 | Ma15-4 | 226.1 | 200 | 45.2 |
23 | HUES6 p26 | Ma15-5 | 238.7 | 200 | 47.7 |
24 | HUES8 p35 | Ma15-6 | 83.4 | 200 | 16.7 |
25 | HUES9 p36 | Ma15-7 | 236.8 | 200 | 47.4 |
26 | ASThiPS4F3 p24 (derived from Astrocytes) | Ma15-8 | 175.9 | 200 | 35.2 |
27 | ASThiPS4F4 p16 (derived from Astrocytes) | Ma15-9 | 164.3 | 200 | 32.9 |
28 | FiPS4F2 p32 (derived from IMR90 fibroblasts) | Ma15-10 | 205.9 | 200 | 41.2 |
29 | FiPS4F5 p29 (derived from IMR90 fibroblasts) | Ma15-11 | 97.6 | 200 | 19.5 |
30 | KiPS4FA p54 (derived from keratinocytes) | Ma15-12 | 129.5 | 200 | 25.9 |
31 | Huv-iPS4F6 p23 (derived from HUVECs) | Ma15-13 | 263.1 | 200 | 52.6 |
32 | IMR90 p6 | Ma15-14 | 76.1 | 200 | 15.2 |
33 | Keratinocytes p3 | Ma15-15 | 138.7 | 200 | 27.7 |
34 | H1 p47 Control | Ma15-16 | 216.4 | 200 | 43.3 |
35 | H1 p47 BMP-4 | Ma15-17 | 137.6 | 200 | 27.5 |
36 | ASThiPS4F5 p12 Control | Ma15-18 | 224.6 | 200 | 44.9 |
37 | ASThiPS4F5 p12 BMP-4 | Ma15-19 | 111.7 | 200 | 22.3 |
38 | Huv-iPS4F6 p23 Control | Ma15-20 | 289.9 | 200 | 58.0 |
39 | Huv-iPS4F6 p23 BMP-4 | Ma15-21 | 172.9 | 200 | 34.6 |
40 | Huv-iPS4F6 p23 ACTIVIN | Ma15-1 | 86.7 | 250 | 21.7 |
Old gDNA of samples have been captured in the previous experiments
- Note:
- Some of gDNA concentrations were re-measured and note that some of them have conc. lower than 50ng/ul, and some of them don't have enough amount for bisulfite conversion
- Samples KERAT MMTA and KiPS4F #8 have high conc., but very little volume. I added ~17ul H2O then re-measured conc. again
- ***I may not do bis-cvt on the two samples, SR-21 and SR22 since the amount of gDNA was too little
No | Sample IDs | Lab IDs | Conc. (ng/ul) |
1 | H9 p43 mTeSR | SR-1 | 62.8 |
2 | H9 p43 ACTIVIN | SR-2 | 90.8 |
3 | H9 p43 BMP4 | SR-3 | 93.7 |
4 | HUV0iPS4F1 p19 | SR-4 | 98.0 |
5 | HUV-hiPS4F1 p20 ACTIVIN | SR-5 | 84.8 |
6 | HUV-hiPS4F1 p20 BMP4 | SR-6 | 78.6 |
7 | HUV-iPS4F3 F19 | SR-7 | 96.9 |
8 | HUV-hiPS4F3 p20 ACTIVIN | SR-8 | 91.9 |
9 | HUV-hiPS4F3 p20 BMP4 | SR-9 | 80.6 |
10 | aMSCs p7 | SR-10 | 184.8 |
11 | MSCiPS #8 p14 (~4 ul) | SR-11 | 105.4 |
12 | MSCiPS #4 p14 | SR-12 | 181.4 |
13 | hNiPS2F p23 | SR-13 | 270.0 |
14 | hKiPS3F-7 p24 (~15ul) | SR-14 | 33.0 |
15 | hNSC | SR-15 | 67.0 |
16 | hFiPS4F-7 p9 | SR-16 | 145.6 |
17 | HUVEC p2 | SR-17 | 245.3 |
18 | hiPS AST4F-4 p9 | SR-18 | 89.9 |
19 | KERAT MMTA | SR-19 | 159.5 |
20 | KiPS4F #8 | SR-20 | 195.7 |
21 | ***olf MSC 19.1ng/ul volume only 3-5ul | SR-21 | 19.1 |
22 | ***KiPS 14ng/ul volume only 3-5ul | SR-22 | 14.0 |
8 samples of bis-cvt gDNA from Dinh
- The list will be filled later and need to re-measure of the conc. with ssDNA Qubit assay
Bisulfite conversion
Reagent prep Add the following reagents to CT Conversion Reagent - H2O 9.0mL - M-Dissolving Buffer 500ul - M-Dilution Buffer 3.0mL Mix at room temperature with frequent vortexing or shaking for 15 minutes Add 144 ml of 100% ethanol to the 36 ml M-Wash Buffer concentrate before use
Lab IDs | Conc. (ng/ul) | Volume for 1200ng | LabID ID | Well IDs | H2O | Total amount (ng) | CT conversion reagent (ul) | Total volume (ul) |
Ap15-1 | 110.0 | 10.91 | Ap15-1 | A1 | 9.09 | 1200.00 | 130.00 | 150.00 |
Ap15-2 | 139.1 | 8.63 | Ap15-2 | B1 | 11.37 | 1200.00 | 130.00 | 150.00 |
Ap15-3 | 145.1 | 8.27 | Ap15-3 | C1 | 11.73 | 1200.00 | 130.00 | 150.00 |
Ap15-4 | 295.5 | 4.06 | Ap15-4 | D1 | 15.94 | 1200.00 | 130.00 | 150.00 |
Ap15-5 | 128.1 | 9.37 | Ap15-5 | E1 | 10.63 | 1200.00 | 130.00 | 150.00 |
Ap15-6 | 153.9 | 7.80 | Ap15-6 | F1 | 12.20 | 1200.00 | 130.00 | 150.00 |
Ap15-7 | 183.5 | 6.54 | Ap15-7 | G1 | 13.46 | 1200.00 | 130.00 | 150.00 |
Ap15-8 | 149.3 | 8.04 | Ap15-8 | H1 | 11.96 | 1200.00 | 130.00 | 150.00 |
Ap15-9 | 111.2 | 10.79 | Ap15-9 | A2 | 9.21 | 1200.00 | 130.00 | 150.00 |
Ap15-10 | 153.5 | 7.82 | Ap15-10 | B2 | 12.18 | 1200.00 | 130.00 | 150.00 |
Ap15-11 | 87.5 | 13.71 | Ap15-11 | C2 | 6.29 | 1200.00 | 130.00 | 150.00 |
Ap15-12 | 109.7 | 10.94 | Ap15-12 | D2 | 9.06 | 1200.00 | 130.00 | 150.00 |
Ap15-13 | 191.3 | 6.27 | Ap15-13 | E2 | 13.73 | 1200.00 | 130.00 | 150.00 |
Ap15-14 | 108.7 | 11.04 | Ap15-14 | F2 | 8.96 | 1200.00 | 130.00 | 150.00 |
Ap15-15 | 134.2 | 8.94 | Ap15-15 | G2 | 11.06 | 1200.00 | 130.00 | 150.00 |
Ap15-16 | 50.2 | 23.90 | Ap15-16 | H2 | -3.90 | 1200.00 | 130.00 | 150.00 |
Ap15-17 | 183.0 | 6.56 | Ap15-17 | A3 | 13.44 | 1200.00 | 130.00 | 150.00 |
Ap15-18 | 82.6 | 14.53 | Ap15-18 | B3 | 5.47 | 1200.00 | 130.00 | 150.00 |
Ap15-19 | 263.4 | 4.56 | Ap15-19 | C3 | 15.44 | 1200.00 | 130.00 | 150.00 |
Ma15-1 | 86.7 | 13.84 | Ma15-1 | D3 | 6.16 | 1200.00 | 130.00 | 150.00 |
Ma15-2 | 215.2 | 5.58 | Ma15-2 | E3 | 14.42 | 1200.00 | 130.00 | 150.00 |
Ma15-3 | 252.0 | 4.76 | Ma15-3 | F3 | 15.24 | 1200.00 | 130.00 | 150.00 |
Ma15-4 | 226.1 | 5.31 | Ma15-4 | G3 | 14.69 | 1200.00 | 130.00 | 150.00 |
Ma15-5 | 238.7 | 5.03 | Ma15-5 | H3 | 14.97 | 1200.00 | 130.00 | 150.00 |
Ma15-6 | 83.4 | 14.39 | Ma15-6 | A4 | 5.61 | 1200.00 | 130.00 | 150.00 |
Ma15-7 | 236.8 | 5.07 | Ma15-7 | B4 | 14.93 | 1200.00 | 130.00 | 150.00 |
Ma15-8 | 175.9 | 6.82 | Ma15-8 | C4 | 13.18 | 1200.00 | 130.00 | 150.00 |
Ma15-9 | 164.3 | 7.30 | Ma15-9 | D4 | 12.70 | 1200.00 | 130.00 | 150.00 |
Ma15-10 | 205.9 | 5.83 | Ma15-10 | E4 | 14.17 | 1200.00 | 130.00 | 150.00 |
Ma15-11 | 97.6 | 12.30 | Ma15-11 | F4 | 7.70 | 1200.00 | 130.00 | 150.00 |
Ma15-12 | 129.5 | 9.27 | Ma15-12 | G4 | 10.73 | 1200.00 | 130.00 | 150.00 |
Ma15-13 | 263.1 | 4.56 | Ma15-13 | H4 | 15.44 | 1200.00 | 130.00 | 150.00 |
Ma15-14 | 76.1 | 15.77 | Ma15-14 | A5 | 4.23 | 1200.00 | 130.00 | 150.00 |
Ma15-15 | 138.7 | 8.65 | Ma15-15 | B5 | 11.35 | 1200.00 | 130.00 | 150.00 |
Ma15-16 | 216.4 | 5.55 | Ma15-16 | C5 | 14.45 | 1200.00 | 130.00 | 150.00 |
Ma15-17 | 137.6 | 8.72 | Ma15-17 | D5 | 11.28 | 1200.00 | 130.00 | 150.00 |
Ma15-18 | 224.6 | 5.34 | Ma15-18 | E5 | 14.66 | 1200.00 | 130.00 | 150.00 |
Ma15-19 | 111.7 | 10.74 | Ma15-19 | F5 | 9.26 | 1200.00 | 130.00 | 150.00 |
Ma15-20 | 289.9 | 4.14 | Ma15-20 | G5 | 15.86 | 1200.00 | 130.00 | 150.00 |
Ma15-21 | 172.9 | 6.94 | Ma15-21 | H5 | 13.06 | 1200.00 | 130.00 | 150.00 |
Program 98°C for 10 minutes 64°C for 2.5 hours 4°C storage up to 20 hours Measure conc. with ssDNA Qubit assay (used 1ul of bis-cvt gDNA)