Alice:LabNotes/2007-11-14: Difference between revisions

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[[Image:ZhangLab_2 2007-11-14 17hr 10min.jpg]]
[[Image:ZhangLab_2 2007-11-14 17hr 10min.jpg]]


Also, I did PCR on sample #14 with newly designed primers rs1264899 along with template GC1EP, GC1F, GM20431, PGP1.
<br>
<b>Experiment 2: PCR on sample #14 with newly designed primer rs1264899 </b> <br>
Summary of reactions:
{| {{table}}
| align="center" style="background:#f0f0f0;"|'''Reaction'''
| align="center" style="background:#f0f0f0;"|'''SNP'''
| align="center" style="background:#f0f0f0;"|'''PCR primer'''
| align="center" style="background:#f0f0f0;"|'''Sample'''
|-
| 1 ||rs1264899||rs1264899_F/rs1264899_R||GC1EP
|-
| 2 ||rs1264899||rs1264899_F/rs1264899_R||GC1F
|-
| 3 ||rs1264899||rs1264899_F/rs1264899_R||GM20432 cDNA
|-
| 4 ||rs1264899||rs1264899_F/rs1264899_R||PGP1 gDNA
|-
|
|}
{| {{table}}
| align="center" style="background:#f0f0f0;"|''''''
| align="center" style="background:#f0f0f0;"|'''one reaction'''
| align="center" style="background:#f0f0f0;"|'''total'''
|-
| Template ||1||17
|-
| 10x Buffer||5||85
|-
| 10mM dNTP||1||17
|-
| 10uM forward primer + reverse primer||1||17
|-
| 50x SYBG I||0.4||6.8
|-
| Jump Start Tag DNA Polymerase||1||17
|-
| H2O||40.6||690.2
|-
|
|}
 
95C 3min -> 35 cycles of (95C 15S -> 58C 30S -> 72C 1min) -> 72C 3min <br>
 
I forgot the reaction was only for 4 samples, therefore the left over master mix is stored in -20 fridge and saved for later use.

Latest revision as of 00:13, 1 December 2007

I ran a gel of the PCR product from repeated experiment on sample #1,2,3,12,14,16. The gel image shows that all the reactions worked fine, so they can be prepared for sequencing. Eventually, the post PCR products are prepared with EXOSap.
File:ZhangLab 2 2007-11-14 17hr 10min.jpg


Experiment 2: PCR on sample #14 with newly designed primer rs1264899
Summary of reactions:

Reaction SNP PCR primer Sample
1 rs1264899 rs1264899_F/rs1264899_R GC1EP
2 rs1264899 rs1264899_F/rs1264899_R GC1F
3 rs1264899 rs1264899_F/rs1264899_R GM20432 cDNA
4 rs1264899 rs1264899_F/rs1264899_R PGP1 gDNA
' one reaction total
Template 1 17
10x Buffer 5 85
10mM dNTP 1 17
10uM forward primer + reverse primer 1 17
50x SYBG I 0.4 6.8
Jump Start Tag DNA Polymerase 1 17
H2O 40.6 690.2

95C 3min -> 35 cycles of (95C 15S -> 58C 30S -> 72C 1min) -> 72C 3min

I forgot the reaction was only for 4 samples, therefore the left over master mix is stored in -20 fridge and saved for later use.