Noi/NOTES/2014-10-3
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Measurement of the size distribution of WGBS libraries generated from cfDNA[edit]
- To verify the size distribution of wGBS libraries generated from cfDNA from plasma samples, I will ask Richard in Palsson's lab to run bioanalyzer. I might include cfDNA for some samples with a high yield (>10ng) to compare.
- Things to do: List samples to be included for Bioanalyzer
- WGBS libraries: Use the High Sensitivity DNA Analysis Kits for 50-7,000bp.
- The sensitivity of the kit is 5pg/ul
- We can run 10 samples/chip at a time.
- Samples to be included: 24 WGBS libraries from normal control and from cancer patients.
- WGBS libraries: Use the High Sensitivity DNA Analysis Kits for 50-7,000bp.
- Today I initially ran 10 samples of those libraries that were used for BSPP capture
- As the library has a tight band, I need to prepare library in the range of 5-500pg to load 1ul. Richard suggested to load 1ul, I then dilute libraries to 1ng/ul in total 5ul. Note that if the size of libraries is wide, we can load as high as 10ng/ul sample.
Sample | Conc. (ng/ul) | Volume to pipette (ul) | H2O (ul) |
NC-6 | 4.25 | 1.18 | 3.82 |
NC-9 | 3.72 | 1.34 | 3.66 |
NC-27 | 6.88 | 0.73 | 4.27 |
NC-30 | 6.46 | 0.77 | 4.23 |
6-P-2 | 12.60 | 0.40 | 4.60 |
6-P-3 | 6.16 | 0.81 | 4.19 |
6-P-4 | 10.40 | 0.48 | 4.52 |
6-P-5 | 5.43 | 0.92 | 4.08 |
cf3 | 9.43 | 0.53 | 4.47 |
cf4 | 9.60 | 0.52 | 4.48 |
- Instruction of Bioanalyzer High Sensitivity DNA Analysis Kits
- We actually can load 11 samples. I then loaded cf4 sample in duplicates.
Result[edit]
- Log file
- From the result, it showed that there is no different of the cfDNA size between samples from normal control and cancer patients.
- One thing that's weird for me is that the library size is ~300bp in 6% TBE which represent the size of cfDNA ~180. However the size of these libraries in Bioanalyzer is only ~205bp.