User:Alexander S Mikheyev/Notebook/rotifer alien genes/2010/05/07

From OpenWetWare
Jump to: navigation, search
OpenWetWare photo.jpg Alien genes Report.pngMain project page
Resultset previous.pngPrevious entry      Next entryResultset next.png


Nuclear Region HS Takara PCR Optimization

Test 4.0ul of 5.0uM Primer Concentration

HS Takara RXN Mixture 3.0ul Instagene conc 1X 5.5 '
Milli Q water 0.1 0.55
Takara HS Buffer 5X 2 11
dNTP 2.5uM 0.8 4.4
Primer Mix 5uM 4
HS Taq 5U 0.1 0.55
Template Instagene 3 16.5
33 6.0ul / RXN
add 4.0ul Primer
A PCR cycle
98C 10 sec
60C 1 min
x35
AMP Lane Template Primer Amt of Template
1 3 Instagene B1 3
2 3 Instagene C1 3
3 3 Instagene D1 3
4 3 Instagene E1 3
5 3 Instagene F1 3
6 4 Instagene B1 3
7 4 Instagene C1 3
8 4 Instagene D1 3
9 4 Instagene E1 3
10 4 Instagene F1 3
11 AV 1ng B1 1
12 AV 1ng C1 1
13 AV 1ng D1 1
14 AV 1ng E1 1
15 AV 1ng F1 1
*AV 2ng Positive Control

Load 2.0ul Sample

06may10B.jpg

AV 2ng - bad dilution

Nuclear Region HS Takara PCR Optimization

Compare 1.0ul of 20uM Primer vs 4.0ul of 5uM (old primer dilution)

HS Takara RXN Mixture conc 1X 3 '
Milli Q water 0.1 0.3
Takara HS Buffer 5X 2 6
dNTP 2.5uM 0.8 2.4
Primer Mix 5uM 4
HS Taq 5U 0.1 0.3
Template Instagene 3 9
18 6.0ul / RXN
add 4.0ul Primer
HS Takara RXN Mixture conc 1X 3
Milli Q water 3.1 9.3
Takara HS Buffer 5X 2 6
dNTP 2.5uM 0.8 2.4
Primer Mix 20uM 1
HS Taq 5U 0.1 0.3
Template Instagene 3 9
27 9.0ul / RXN
add 1.0ul Primer
A PCR cycle
98C 10 sec
60C 1 min
x35
AMP Lane Template Primer Amt of Template Primer
1 3 Instagene F2 3 4.0ul of 5uM
2 4 Instagene F2 3 4.0ul of 5uM
3 AV 20ng F2 1 4.0ul of 5uM
4 3 Instagene F2 3 1.0ul of 20uM
5 4 Instagene F2 3 1.0ul of 20uM
6 AV 20ng F2 1 1.0ul of 20uM
*AV 20ng Positive Control

Load 2.0ul Sample

07May10B.jpg

well 2 has a weak band