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= CRC oligo pool from Illumina = [[noi:DMR220k_LabNotes|'''Calendar''']] * I received one tube of CRC (colorectal cancer) oligo pool from Xuyu. The total volume of oligo ~1mL * The concentration of oligo mix is 100uM and there are 1,024 oligos in this pool. * I verify concentration by Qubit ssDNA assay by measuring non-diluted and 10X, 50X and 100X diluted oligo in TE Buffer. :{| {{table}} border = 1 | align="center" style="width:120px;background:#f0f0f0;"|'''Sample''' | align="center" style="width:80px;background:#f0f0f0;"|'''Conc. in the Qubit''' | align="center" style="width:60px;background:#f0f0f0;"|'''Unit''' | align="center" style="width:60px;background:#f0f0f0;"|'''uL used''' | align="center" style="width:60px;background:#f0f0f0;"|'''Dilution''' | align="center" style="width:80px;background:#f0f0f0;"|'''Sample Conc. (ng/ul)''' | align="center" style="width:80px;background:#f0f0f0;"|'''Conc. of original tube (ng/ul)''' |- | 1ul non-diluted||Out Of Range||||1|||||| |- | 2ul non-diluted||Out Of Range||||2|||||| |- | 2ul 10X dilution||Out Of Range||||2|||||| |- | 2ul 100X dilution||140||ng/mL||2||100||14||1,400 |- | 2ul 50X dilution||270||ng/mL||2||100||27||1,350 |} * I also verify the integrity of oligos in 6% TBU gel to see if the level of background is high. [[File:ZhangLab_2 2014-07-10 17hr 22min_CRC-oligos.jpg|350px]] 1 = 1ul of non-diluted CRC oligos 2 = 2ul of non-diluted CRC oligos 3 = 2ul of 10X diluted CRC oligos 4 = 2ul of 100X diluted CRC oligos 5 = 2ul of 50X diluted CRC oligos * It looked like there is very high background of truncated oligos as expect. I may need to do size selection of this oligo pool (~12ug) before capture to avoid any background during capture. In addition, I have a lot of oligos (at least 1,000ug) for this experiment. ==== PAGE size selection of CRC oligos ==== * I mix 3ug of CRC oligo in total 100ul of TBU loading dye * For 4 gel I mixed 8.57ul of 1.4ug/ul CRC oligo with 192ul H2O and 200ul 2X TBU loading dye and split 100ul in 200ul PCR tube. [[File:ZhangLab_2 2014-07-11 12hr 02min_CRC_PAGE-SS_1.jpg| 350px]] [[File:ZhangLab_2 2014-07-11 12hr 02min_CRC_PAGE-SS_2.jpg|350px]] [[File:ZhangLab_2 2014-07-11 12hr 02min_CRC_PAGE-SS_3.jpg| 350px]] [[File:ZhangLab_2 2014-07-11 12hr 02min_CRC_PAGE-SS_4.jpg| 350px]] :- Precipitate DNA in 12 LoBind tubes :- Resuspend each tube with 20ul --> total 240ul :- Quantify concentration with Qubit ssDNA assay --> conc. = XX ng/ul :- Verify purified oligos in 6% TBU gel
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