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Daniel:Notebook/HiResChrPaint/2014-9-10
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=Padlock Probes II= [[Daniel:Notebook/HiResChrPaint|Back to Calendar]] ===Nanodrop Results=== {| class="wikitable" <hiddentext>generated with [[:de:Wikipedia:Helferlein/VBA-Macro for EXCEL tableconversion]] V1.8</hiddentext> |- style="background-color:#B8CCE4;font-size:12pt;font-weight:bold" | width="65" height="30" valign="bottom" | Sample | width="65" align="center" | 1:1 ng/uL | width="65" align="center" | 5:1 ng/uL | width="65" align="center" | 5:1 ng/uL Projected | width="65" align="center" | 10:1 ng/uL | width="65" align="center" | 10:1 ng/uL Projected | width="65" align="center" | Average ng/uL | width="65" align="center" | Standard Deviation | width="65" align="center" | ug in 100 uL |- style="font-size:12pt" |style="font-weight:bold" height="15" valign="bottom" | PPRL-A | align="center" align="center" valign="bottom" | 1944.1 | align="center" align="center" valign="bottom" | 373 | align="center" align="center" valign="bottom" | 1865 | align="center" align="center" valign="bottom" | 184.4 | align="center" align="center" valign="bottom" | 1844 | align="center" align="center" valign="bottom" | 1884.4 | align="center" align="center" valign="bottom" | 52.8 |style="font-weight:bold" align="center" align="center" valign="bottom" | 188.4 |- style="background-color:#D9D9D9;font-size:12pt" |style="font-weight:bold" height="15" valign="bottom" | PPRL-B | align="center" align="center" valign="bottom" | 2064.8 | align="center" align="center" valign="bottom" | 421.7 | align="center" align="center" valign="bottom" | 2108.5 | align="center" align="center" valign="bottom" | 203 | align="center" align="center" valign="bottom" | 2030 | align="center" align="center" valign="bottom" | 2067.8 | align="center" align="center" valign="bottom" | 39.3 |style="font-weight:bold" align="center" align="center" valign="bottom" | 206.8 |} Usually after ethanol precipitation I get really high results, so I'll just say these are fine and continue as though I have a reasonable amount of material. ==DpnII/USER Digestion== '''Reaction Table''' {| class="wikitable" <hiddentext>generated with [[:de:Wikipedia:Helferlein/VBA-Macro for EXCEL tableconversion]] V1.8</hiddentext> |- style="background-color:#B8CCE4;font-size:12pt;font-weight:bold" align="center" | width="65" height="30" | Reagent | width="71" | PPRL-A1 (uL) | width="71" | PPRL-A2 (uL) | width="71" | PPRL-B1 (uL) | width="65" | PPRL-B2 (uL) |- style="font-size:12pt" |style="background-color:#FCD5B4" height="15" valign="bottom" | amplicon | align="center" align="center" valign="bottom" | 50 | align="center" align="center" valign="bottom" | 50 | align="center" align="center" valign="bottom" | 50 | align="center" align="center" valign="bottom" | 50 |- style="font-size:12pt" |style="background-color:#FCD5B4" height="15" valign="bottom" | ug Input | align="center" align="center" valign="bottom" | 94.2 | align="center" align="center" valign="bottom" | 94.2 | align="center" align="center" valign="bottom" | 103.4 | align="center" align="center" valign="bottom" | 103.4 |- style="font-size:12pt" |style="background-color:#FCD5B4" height="15" valign="bottom" | USER | align="center" align="center" valign="bottom" | 5 | align="center" align="center" valign="bottom" | 5 | align="center" align="center" valign="bottom" | 5 | align="center" align="center" valign="bottom" | 5 |- style="font-size:12pt" |style="background-color:#FCD5B4" height="15" valign="bottom" | nf H2O | align="center" align="center" valign="bottom" | 30 | align="center" align="center" valign="bottom" | 30 | align="center" align="center" valign="bottom" | 30 | align="center" align="center" valign="bottom" | 30 |- style="font-size:12pt;font-weight:bold" |style="background-color:#FCD5B4" height="15" valign="bottom" | Total | align="center" align="center" valign="bottom" | 85 | align="center" align="center" valign="bottom" | 85 | align="center" align="center" valign="bottom" | 85 | align="center" align="center" valign="bottom" | 85 |} #Set up reaction according to table in a 0.2 mL PCR tube #Incubate for 2 hours at 37C #Add 10 uL DpnII buffer (10X) #Incubate at 94C for 2 minutes #Cool to 37C and incubate for 3 minutes #Add 5 uL DpnII and incubate at 37C for 2 hours #Heat inactivate DpnII at 65C for 20 minutes #[[Daniel:Protocols/ ===Nanodrop Results=== {| class="wikitable" <hiddentext>generated with [[:de:Wikipedia:Helferlein/VBA-Macro for EXCEL tableconversion]] V1.8</hiddentext> |- style="background-color:#B8CCE4;font-size:12pt;font-weight:bold" | width="65" height="15" valign="bottom" | Sample | width="65" align="center" valign="bottom" | PPRL-A1 | width="65" align="center" valign="bottom" | PPRL-A2 | width="65" align="center" valign="bottom" | PPRL-B1 | width="65" align="center" valign="bottom" | PPRL-B2 |- style="font-size:12pt" |style="font-weight:bold" height="15" valign="bottom" | ng/uL | align="center" align="center" valign="bottom" | 154.1 | align="center" align="center" valign="bottom" | 131 | align="center" align="center" valign="bottom" | 153 | align="center" align="center" valign="bottom" | 163.7 |- style="background-color:#D9D9D9;font-size:12pt" |style="font-weight:bold" height="15" valign="bottom" | ug in 40 uL | align="center" align="center" valign="bottom" | 6.2 | align="center" align="center" valign="bottom" | 5.2 | align="center" align="center" valign="bottom" | 6.1 | align="center" align="center" valign="bottom" | 6.5 |- style="font-size:12pt" |style="font-weight:bold" height="15" valign="bottom" | ug total | align="center" align="center" valign="bottom" | | align="center" align="center" valign="bottom" | | align="center" align="center" valign="bottom" | |style="font-weight:bold" align="center" align="center" valign="bottom" | 24.1 |} ===TBE Gel=== #For each sample, 5 uL TBE, 2 uL 6x dye, 1 uL sample; Load 7.5 uL each #Run gel at 220V for 25 minutes {{Template:GelLanes|Lane2=Low Mass Ladder|Lane3=PPRL-A, pre-digest 10:1|Lane4=PPRL-A, pre-digest 40:1|Lane5=PPRL-A1|Lane6=PPRL-A2|Lane7=PPRL-B, pre-digest 10:1|Lane8=PPRL-B, pre-digest 40:1|Lane9=PPRL-B1|Lane10=PPRL-B2}} '''Gel Image''' [[Image:2014-09-09-PPRL.jpg|800px]] [[Category:HRCP]] [[Category:DpnII]] [[Category:USER]] [[Category:TBE Gel]] [[Category:20140909]]
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