User:Alexander S Mikheyev/Notebook/rotifer alien genes/2010/04/12

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Ethanol Post-PCR Clean Up


1) For each 10 ul of PCR product, prepare the following mixture in strip-cap tube. 1 ul of 3M sodium acetate (NaOAc) pH 5.2 25 ul chilled absolute ethanol. 2) Transfer the PCR product (20ul) into the tube containing the mixture. 3) Vortex the tube and then store it at -20 for 30-50 min. 4) Spin for 30 min at maximum speed 14.000 rpm. 5) Rremove the supernatant carefully and don't disturb the pellet. 6) Wash the pellet with 70% ethanol. And spin the tube at maximum speed for 5 min. 7) Remove the supernatant and then dry the pellet – air dry. 8) Resuspend the pellet in 10 ul in sterile water.

Image:14April10 Plate.jpg

Ethanol Clean # MIX Contents ' Size Gene CCA3 Result Ethanol Cleaned ng/ul Sample Type Total Product Remaining bp/ng ul to mix ng added ng/ul
123AV10004 FWDAV10007 RVS8691AV212 Agood 04Mar10495.3DNAx 9ul0.056989989644461.14395581149371566.601313432836
21AV10007 FWDAV10011 RVS9262AV212 Agood 04Mar10437.4DNAx 9ul0.0472252213344851.38049177978421603.827104477612
32AV10011 FWDAV10016 RVS11924AV212 Agood 02Mar10314.9DNAx 9ul0.0264089231801412.4686364304233777.373611940299
424AV10016 FWDAV10020 RVS7020AV212 Agood 04Mar10408.3DNAx 9ul0.0581623931623931.12089661903561457.662089552239
525AV10020 FWDAV10025 RVS11202AV212 Agood 04Mar10403.1DNAx 9ul0.0359846455991.81171799153575730.30352238806
64AV10025 FWDAV10028 RVS6843AV212 Agood 04Mar10330.8DNAx 9ul0.0483413707438261.34861773358119446.122746268657
734AV10029 FWDAV10034 RVS12093AV240 Bgood 08Mar10380.5DNAx 9ul0.0314644835855452.07198791848903788.391402985075
840AV10034 FWDAV10039 RVS14180AV240 Bgood 18Mar101128.6DNAx 9ul0.0795909732016920.81911336459648924.451343283582
950AV10039 FWDAV10041 FWD19555AV240 Bgood 18Mar101888.1DNAx 9ul0.0965533111736130.675212782019671274.86925373134
1030EU643474 FWDEU643474 RVS10062AV240 Bgood 04Mar10206DNAx 9ul0.0204730669846953.18438023474859655.982328358209
1131AV10045 FWDAV10047 RVS6762AV240 Bgood 04Mar10393.3DNAx 9ul0.0581632653061221.12087981146897440.842029850746
1232AV10047 FWDAV10049 RVS12719AV240 Bgood 04Mar10423.1DNAx 9ul0.0332651938045441.95982714647044829.202865671642
1333AV10049 FWDAV10053 RVS9353AV240 Bgood 04Mar10364.3DNAx 9ul0.0389500694964181.67378468623121609.75976119403
25138AV10083 FWDAV10084.2 RVS5400AV TEL L Agood 05Apr10214.1DNAx 7.5ul0.0396481481481481.64431462491373352.04776119403
1478AV10091 FWDAV10092.2 RVS8127AV TEL L Agood 19Mar10515.5DNAx 9ul0.0634305401747261.02780190222505529.831880597015
1585AV10092.2 FWDAV10094 RVS7036AV TEL L Agood 01Apr1353.9DNAx 9ul0.192424673109720.33880286138552458.705194029851
1616AV10094 FWDAV10098 RVS8600AV TEL L Agood 02Mar10334.4DNAx 9ul0.0388837209302331.67664071984575560.668656716418
17134AV10098 FWDAV10104 FWD8000AV TEL L A&Bgood 01Apr466.4DNAx 9ul0.05831.11825094083613521.55223880597
1851AV10104 RVSAV10106 RVS9618AV_TEL_L_Bgood 16Mar430.3DNAx 9ul0.0447390309835721.45720701627813627.036179104478
19111AV10106 FWDAV10108.2 RVS13400AV_TEL_L_Bgood 29Mar218.4DNAx 9ul0.0162985074626874873.6
20112AV10107.1 FWDAV10108 RVS7900AV TEL L Bgood 29Mar214.1DNAx 9ul0.0271012658227852.40557139570713515.032835820896
2155AV10108 FWDAV10111 RVS14846AV_TEL_L_Bgood 16Mar210.7DNAx 9ul0.0141923750505194.5935954777607967.870567164179
22117AV10111 FWDAV10113 RVS8400AV TEL L Bgood 29Mar355.8DNAx 9ul0.0423571428571431.5391507890564547.629850746269
23128AV10113 FWDTEL_L_B_3 RVS11500AV TEL L Bgood 01Apr422.7DNAx 9ul0.036756521739131.77367244685021749.731343283582
24131AV10114 FWDTEL_L_B_4 RVS8500AV TEL L Bgood 01Apr374.2DNAx 9ul0.0440235294117651.48089057651348554.149253731343

Alien Genes New Primer Screen

Reaction mixture conc. 1x x16.5 ' lane '
Temp??227Ithaca 2 A3
TopTaq Buffer10x116.528Ithaca 2 A4
dNTP mix10 mM each0.23.329Ithaca 2 A5
Primer mix (histone_F1/histone_R1)5 uM each116.537Tsukuba 3 D2
TopTaq5 U/ul0.050.82540Woods Hole 1 A6
Milli Q5.7594.87543Yokohama B5
10 ul47Harvard A1
48Ithaca 2 A6
PCR cycle50Tsukuba 1 B5
94C3 min51Yokohama A2
55Yokohama B2
98C10 sec56Yokohama A4
60C3 min61Pennsylvania(PA) B3
x3562Ithaca1 C3
75Baltimore A5
72C5 min76Pennsylvania (PA) B1


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