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== Bisulfite conversion == * I did bisulfite converssion by usingEZ DNA Methylation-Lightning™ Kit -Zymo and followed the protocol of the kit === Bisulfite conversion set up === {| {{table}} class = wikitable | align="center" style="background:#f0f0f0;"|'''Samples''' | align="center" style="background:#f0f0f0;"|'''Conc. (ng/ul)''' | align="center" style="background:#f0f0f0;"|'''Volume for 600ng (ul)''' | align="center" style="background:#f0f0f0;"|'''Go to tube''' | align="center" style="background:#f0f0f0;"|'''H2O''' | align="center" style="background:#f0f0f0;"|'''Lightning Conv Reagent''' |- | CANC_1||47.70||12.58||MN#1||7.42||130.00 |- | CANC_2||45.40||13.22||MN#2||6.78||130.00 |- | CANC_3||41.40||14.49||MN#3||5.51||130.00 |- | CANC_4||47.10||12.74||MN#4||7.26||130.00 |- | CANC_5||68.20||8.80||MN#5||11.20||130.00 |- | SR-3||62.20||9.65||MN#6||10.35||130.00 |- | MA-16||85.00||7.06||MN#7||12.94||130.00 |- | MA-17||49.40||12.15||MN#8||7.85||130.00 |- | F10||106.00||5.66||MN#9||14.34||130.00 |- | G10||108.00||5.56||MN#10||14.44||130.00 |- | G11||82.60||7.26||MN#11||12.74||130.00 |- | NTC||0||0||MN#12||20.00||130.00 |} '''Program''' 1. 98°C for 8 minutes 2. 54°C for 60 minutes 3. 4°C hold * I elute with Elution buffer 16ul * I used 1ul for ssDNA Qubit assay. The volume left after assay ~13ul. ==== Qubit ss DNA assay ==== * Dilute 200X (add 1ul of bis-cvt DNA in total 200 assay volume {| {{table}} class = wikitable | align="center" style="background:#f0f0f0;"|'''Sample''' | align="center" style="background:#f0f0f0;"|'''Conc. in the Qubit''' | align="center" style="background:#f0f0f0;"|'''''' | align="center" style="background:#f0f0f0;"|'''uL used''' | align="center" style="background:#f0f0f0;"|'''Dilution''' | align="center" style="background:#f0f0f0;"|'''Sample conc.''' | align="center" style="background:#f0f0f0;"|'''''' | align="center" style="background:#f0f0f0;"|'''Yields in 13ul (ng)''' | align="center" style="background:#f0f0f0;"|'''% recovery''' |- | MN_1||175||ng/mL||1||200||34.9||ng/ul||453.7||75.62 |- | MN_2||122||ng/mL||1||200||24.5||ug/mL||318.5||53.08 |- | MN_3||122||ng/mL||1||200||24.5||ug/mL||318.5||53.08 |- | MN_4||87.6||ng/mL||1||200||17.5||ug/mL||227.5||37.92 |- | MN_5||108||ng/mL||1||200||21.7||ug/mL||282.1||47.02 |- | MN_6||131||ng/mL||1||200||26.2||ug/mL||340.6||56.77 |- | MN_7||107||ng/mL||1||200||21.4||ug/mL||278.2||46.37 |- | MN_8||71.4||ng/mL||1||200||14.3||ug/mL||185.9||30.98 |- | MN_9||61.5||ng/mL||1||200||12.3||ug/mL||159.9||26.65 |- | MN_10||173||ng/mL||1||200||34.6||ug/mL||449.8||74.97 |- | MN_11||72.9||ng/mL||1||200||14.6||ug/mL||189.8||31.63 |}
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