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== Sequencing library concentration by Nanodrop == * Note: To combine equal molecular ratio of sequencing libraries, I measured concentration of each library by Nanodrop. The corresponding concentration may not be very accurate as PQ (at specific 300-600bp range) and the concentration may be overestimated. However, this should be good enough. For some libraries which have very low concentration (<5.5ng/ul), I add maximum volume 35ul to the library pool to save half of them for DSN nomalization. {| class="wikitable" style="text-align:center; | align="center" style="background:#f0f0f0;"|'''Sample ID''' | align="center" style="background:#f0f0f0;"|'''ng/ul''' | align="center" style="background:#f0f0f0;"|'''Yields in 70ul (ng)''' | align="center" style="background:#f0f0f0;"|'''Volume for 200ng (ul)''' | align="center" style="background:#f0f0f0;"|'''Total amount (ng)<br>pooled in the seqlib''' | align="center" style="background:#f0f0f0;"|'''Approximated volume left (ul)''' |- | a3||10.51||735.70||19.03||200.00||50.97 |- | b3||12.93||905.10||15.47||200.00||54.53 |- | c3||7.86||550.20||25.45||200.00||44.55 |- | d3||6.56||459.20||30.49||200.00||39.51 |- | f3||12.22||855.40||16.37||200.00||53.63 |- | f3||12.60||882.00||15.87||200.00||54.13 |- | g3||5.48||383.60||35.00||191.80||35.00 |- | h3||8.37||585.90||23.89||200.00||46.11 |- | a4||10.34||723.80||19.34||200.00||50.66 |- | b4||16.43||1150.10||12.17||200.00||57.83 |- | c4||40.51||2835.70||4.94||200.00||65.06 |- | d4||30.31||2121.70||6.60||200.00||63.40 |- | e4||20.63||1444.10||9.69||200.00||60.31 |- | f4||22.15||1550.50||9.03||200.00||60.97 |- | g4||15.48||1083.60||12.92||200.00||57.08 |- | h4||13.09||916.30||15.28||200.00||54.72 |- | a5||13.60||952.00||14.71||200.00||55.29 |- | b5||29.56||2069.20||6.77||200.00||63.23 |- | c5||15.63||1094.10||12.80||200.00||57.20 |- | d5||30.58||2140.60||6.54||200.00||63.46 |- | e5||29.43||2060.10||6.80||200.00||63.20 |- | f5||16.55||1158.50||12.08||200.00||57.92 |- | g5||11.05||773.50||18.10||200.00||51.90 |- | h5||12.59||881.30||15.89||200.00||54.11 |- | a6||12.06||844.20||16.58||200.00||53.42 |- | b6||18.89||1322.30||10.59||200.00||59.41 |- | c6||12.57||879.90||15.91||200.00||54.09 |- | d6||13.98||978.60||14.31||200.00||55.69 |- | e6||18.46||1292.20||10.83||200.00||59.17 |- | f6||18.98||1328.60||10.54||200.00||59.46 |- | g6||12.28||859.60||16.29||200.00||53.71 |- | h6||4.43||310.10||35.00||155.05||35.00 |- | a7||9.23||646.10||21.67||200.00||48.33 |- | b7||20.87||1460.90||9.58||200.00||60.42 |- | c7||19.38||1356.60||10.32||200.00||59.68 |- | d7||33.00||2310.00||6.06||200.00||63.94 |- | e7||16.55||1158.50||12.08||200.00||57.92 |- | f7||16.68||1167.60||11.99||200.00||58.01 |- | g7||13.04||912.80||15.34||200.00||54.66 |- | h7||9.28||649.60||21.55||200.00||48.45 |- | a8||5.89||412.30||33.96||200.00||36.04 |- | b8||9.28||649.60||21.55||200.00||48.45 |- | c8||10.88||761.60||18.38||200.00||51.62 |- | d8||10.91||763.70||18.33||200.00||51.67 |- | e8||10.18||712.60||19.65||200.00||50.35 |- | f8||10.30||721.00||19.42||200.00||50.58 |- | g8||3.65||255.50||35.00||127.75||35.00 |- | h8||4.04||282.80||35.00||141.40||35.00 |- | a9||5.71||399.70||35.03||200.00||34.97 |- | b9||7.70||539.00||25.97||200.00||44.03 |- | c9||13.63||954.10||14.67||200.00||55.33 |- | d9||15.94||1115.80||12.55||200.00||57.45 |- | e9||10.19||713.30||19.63||200.00||50.37 |- | f9||14.79||1035.30||13.52||200.00||56.48 |- | g9||14.80||1036.00||13.51||200.00||56.49 |- | h9||10.99||769.30||18.20||200.00||51.80 |- | a10||4.78||334.60||35.00||167.30||35.00 |- | b10||10.83||758.10||18.47||200.00||51.53 |- | c10||7.88||551.60||25.38||200.00||44.62 |- | d10||13.33||933.10||15.00||200.00||55.00 |- | e10||8.53||597.10||23.45||200.00||46.55 |- | f10||10.81||756.70||18.50||200.00||51.50 |- | g10||10.17||711.90||19.67||200.00||50.33 |- | h10||6.92||484.40||28.90||200.00||41.10 |- | a11||6.90||483.00||28.99||200.00||41.01 |- | b11||9.62||673.40||20.79||200.00||49.21 |- | c11||8.67||606.90||23.07||200.00||46.93 |- | d11||12.25||857.50||16.33||200.00||53.67 |- | e11||11.38||796.60||17.57||200.00||52.43 |- | f11||11.91||833.70||16.79||200.00||53.21 |- | g11||12.44||870.80||16.08||200.00||53.92 |- | h11||9.89||692.30||20.22||200.00||49.78 |} * Performed PAGE size-selection (200ng each from 72 libraries, in 4 of 5-well gels) [[File:ZhangLab_2 2013-02-12 12hr 21min_PAGE-ss1.jpg| 450px]] [[File:ZhangLab_2 2013-02-12 12hr 24min_PAGE-ss2.jpg| 480px]] * Resuspend ~70ul with H20 * Verify the size in 6% TBE gel [[File:ZhangLab_2 2013-02-12 15hr 59min.jpg| 240px]] * Quantify concemtration by Qubit dsDNA HS assay ** Average conc. of = 19.73ng/ul ** Conc. based on MW (size ~400bp) 243,228g/mol = 81.1nM * '''Library ID: NP-SkinMB_N2_Ind17-88_Feb07.2013'''. The libraires were sequenced in HL144 run (lane1), HL145 (lane ) * For sample tracking, please refer to the table on: http://genome-tech.ucsd.edu/LabNotes/index.php/Noi/NOTES/2013-1-31 * The plate labeled: ''' SkinMB.Ind17-88.Feb07,2013''' {| class="wikitable" style="text-align:center; | align="center" style="background:#f0f0f0;"|'''Position in 96-well''' | align="center" style="background:#f0f0f0;"|'''Indx''' | align="center" style="background:#f0f0f0;"|'''Position in 96-well''' | align="center" style="background:#f0f0f0;"|'''Indx''' |- | A3||PCR_R.N2Indx17||E7||PCR_R.N2Indx53 |- | B3||PCR_R.N2Indx18||F7||PCR_R.N2Indx54 |- | C3||PCR_R.N2Indx19||G7||PCR_R.N2Indx55 |- | D3||PCR_R.N2Indx20||H7||PCR_R.N2Indx56 |- | E3||PCR_R.N2Indx21||A8||PCR_R.N2Indx57 |- | F3||PCR_R.N2Indx22||B8||PCR_R.N2Indx58 |- | G3||PCR_R.N2Indx23||C8||PCR_R.N2Indx59 |- | H3||PCR_R.N2Indx24||D8||PCR_R.N2Indx60 |- | A4||PCR_R.N2Indx25||E8||PCR_R.N2Indx61 |- | B4||PCR_R.N2Indx26||F8||PCR_R.N2Indx62 |- | C4||PCR_R.N2Indx27||G8||PCR_R.N2Indx63 |- | D4||PCR_R.N2Indx28||H8||PCR_R.N2Indx64 |- | E4||PCR_R.N2Indx29||A9||PCR_R.N2Indx65 |- | F4||PCR_R.N2Indx30||B9||PCR_R.N2Indx66 |- | G4||PCR_R.N2Indx31||C9||PCR_R.N2Indx67 |- | H4||PCR_R.N2Indx32||D9||PCR_R.N2Indx68 |- | A5||PCR_R.N2Indx33||E9||PCR_R.N2Indx69 |- | B5||PCR_R.N2Indx34||F9||PCR_R.N2Indx70 |- | C5||PCR_R.N2Indx35||G9||PCR_R.N2Indx71 |- | D5||PCR_R.N2Indx36||H9||PCR_R.N2Indx72 |- | E5||PCR_R.N2Indx37||A10||PCR_R.N2Indx73 |- | F5||PCR_R.N2Indx38||B10||PCR_R.N2Indx74 |- | G5||PCR_R.N2Indx39||C10||PCR_R.N2Indx75 |- | H5||PCR_R.N2Indx40||D10||PCR_R.N2Indx76 |- | A6||PCR_R.N2Indx41||E10||PCR_R.N2Indx77 |- | B6||PCR_R.N2Indx42||F10||PCR_R.N2Indx78 |- | C6||PCR_R.N2Indx43||G10||PCR_R.N2Indx79 |- | D6||PCR_R.N2Indx44||H10||PCR_R.N2Indx80 |- | E6||PCR_R.N2Indx45||A11||PCR_R.N2Indx81 |- | F6||PCR_R.N2Indx46||B11||PCR_R.N2Indx82 |- | G6||PCR_R.N2Indx47||C11||PCR_R.N2Indx83 |- | H6||PCR_R.N2Indx48||D11||PCR_R.N2Indx84 |- | A7||PCR_R.N2Indx49||E11||PCR_R.N2Indx85 |- | B7||PCR_R.N2Indx50||F11||PCR_R.N2Indx86 |- | C7||PCR_R.N2Indx51||G11||PCR_R.N2Indx87 |- | D7||PCR_R.N2Indx52||H11||PCR_R.N2Indx88 |} * Continued on : [[http://genome-tech.ucsd.edu/LabNotes/index.php/Noi/NOTES/2013-3-20]], pooled the top 20 libraries for HiSeq sequencing
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