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===Results=== *total RNA. {| {{table}} | align="center" style="background:#f0f0f0;"|'''''' | align="center" style="background:#f0f0f0;"|'''5406.090161''' | align="center" style="background:#f0f0f0;"|'''5''' | align="center" style="background:#f0f0f0;"|'''''' |- | ||3730.570111||2.5||in 10 uL |- | 1. i5XX-i7XX, T7-nXTv2_i5-1, 5X tn5-059, 500 cells rep1||17614.20052||23.21540477||232.1540477 |- | 2. i5XX-i7XX, T7-nXTv2_i5-1, 5X tn5-059, 500 cells rep2||16680.7305||21.82259836||218.2259836 |- | 3. i5XX-i7XX, T7-nXTv2_i5-1, 5X tn5-059, 6 ng pure DNA||142250.0542||209.1813288||2091.813288 |- | 4. i5XX-i7XX, T7-nXTv2_i5-1, 5X tn5-059, NTC||708.0800211||-2.009779054||-20.09779054 |- | |} *TBU gel after IVT, Guanidine HCl to remove proteins *yields from 500 cells approx. half that seen before, will try PAGE purified transposon instead, and v2 version. [[File:ZhangLab 2 2015-08-28 11hr 27min-labeled.jpg|600px]] *qPCR curves *do not gain much info from efficiency based on PCR curves for short versus long primers. seems 502 works better. [[File:2015-09-09 nXTv2 validation, primers testing.bmp|600px]] *after adding barcodes, TBE gel *high amount of primer dimers, maybe since ran at 59C for annealing instead of 63C. next time will try 63C. [[File:ZhangLab 2 2015-09-09 18hr 14min-labeled.jpg|600px]] *gel size selection amounts {| {{table}} | align="center" style="background:#f0f0f0;"|'''''' | align="center" style="background:#f0f0f0;"|'''6007.810179''' | align="center" style="background:#f0f0f0;"|'''2.5''' | align="center" style="background:#f0f0f0;"|'''for 150 ng''' | align="center" style="background:#f0f0f0;"|'''X2''' |- | 21. i502_shrt-i701_long, 8/27/2015, 1. T7-nXTv2_i5-1, 5X tn5-059, 500 cells rep1||23864.16071||23.11343865||6.48973103||12.97946206 |- | 22. i502_shrt-i702_long, 8/27/2015, 2. T7-nXTv2_i5-1, 5X tn5-059, 500 cells rep2||20633.37061||19.38380233||7.738419813||15.47683963 |- | |} *gel size selection [[File:ZhangLab 2 2015-09-10 14hr 03min-labeled.jpg|600px]] *gel size selection validation [[File:ZhangLab 2 2015-09-10 18hr 23min-labeled.jpg|600px]]
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