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====1.2) STD RRBS ''Msp''I digestion set up ==== :{| {{table}} class = wikitable | align="center" style="width:100px;background:#f0f0f0;"|'''CCT-100ng_1''' | align="center" style="width:100px;background:#f0f0f0;"|'''CTT-100ng_2''' | align="center" style="width:100px;background:#f0f0f0;"|'''CTT-F-100ng_1''' | align="center" style="width:100px;background:#f0f0f0;"|'''CTT-F-100ng_2''' | align="center" style="width:100px;background:#f0f0f0;"|'''100ng NTC''' |- |} :{| {{table}} class = wikitable | align="center" style="width:120px;background:#f0f0f0;"|'''Sample''' | align="center" style="width:80px;background:#f0f0f0;"|'''Conc. (ng/ul)''' | align="center" style="width:80px;background:#f0f0f0;"|'''Volume for 100ng (ul)''' | align="center" style="width:80px;background:#f0f0f0;"|'''10X Tango Fuffer''' | align="center" style="width:80px;background:#f0f0f0;"|'''MspI (10U/ul)''' | align="center" style="width:80px;background:#f0f0f0;"|'''H2O to adjust volume (ul)''' | align="center" style="width:80px;background:#f0f0f0;"|'''1ng/ul unmeth-lambda DNA''' | align="center" style="width:80px;background:#f0f0f0;"|'''H2O in MspI mix(ul)''' | align="center" style="width:80px;background:#f0f0f0;"|'''Total (ul)''' |- | 1.1 CTT-frozen||52.30||1.91||2.00||1.00||5.09||0.50||7.50||18.00 |- | 1.2 CTT-FFPE||64.80||1.54||2.00||1.00||5.46||0.50||7.50||18.00 |- | 1.3 NTC||0.00||0.00||2.00||1.00||7.00||0.50||7.50||18.00 |} :<u>'''''Msp''I reaction mix II'''</u> :{| {{table}} border = 1 | align="center" style="width:150px;background:#f0f0f0;"|'''Components''' | align="center" style="width:90px;background:#f0f0f0;"|'''1 rxn''' | align="center" style="width:90px;background:#f0f0f0;"|'''5.5 rxn mix''' |- | MspI (10U/ul)||1.00||5.50 |- | 10X Tango Buffer||2.00||11.00 |- | 1ng/ul unmeth-lambda DNA||0.5||2.75 |- | H2O||7.50||41.25 |- | Total||11.00|| |} :- Add H2O to adjust volume as table above :- Aliquot 11ul to each tube :- Add CTT DNA to each tube following the number in table above :- Mix by gentle pulse-vortexting for 10x (put reaction tube on PCR rack) and spin down --------------------------------------------------------------------------- :- Incubate at 37C for 3hr :- Heat inactivate at 65C for 20min
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