SBB09Ntbk-Jong H. Shin: Difference between revisions
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Then, I followed the instruction below.<br> | Then, I followed the instruction below.<br> | ||
[http://openwetware.org/wiki/Template:SBB-Protocols_PCR2 Cloning by PCR]<br> | [http://openwetware.org/wiki/Template:SBB-Protocols_PCR2 Cloning by PCR]<br> | ||
Since the size is over 4000K, I put the PCR tube in 8k55. | Since the size is over 4000K, I put the PCR tube in 8k55.<br> | ||
<br> | <br> | ||
'''Feb 23 2009'''<br> | '''Feb 23 2009'''<br> | ||
| Line 26: | Line 26: | ||
<br>mine is the 5th one<br> | <br>mine is the 5th one<br> | ||
The size is between 1.5k and 1k; it's about the right size I've expected. Product? Yes.<br> | The size is between 1.5k and 1k; it's about the right size I've expected. Product? Yes.<br> | ||
[http://openwetware.org/images/f/f2/IMG_1608.JPG The picture link for the second product]<br> | |||
mine is the 2nd one<br> | mine is the 2nd one<br> | ||
The result came out the same as the first one. Product? Yes!<br> | The result came out the same as the first one. Product? Yes!<br> | ||
7. Zymo cleanup<br> | 7. Zymo cleanup<br><br> | ||
'''Feb 25 2009'''<br> | |||
---- | |||
Digestion<br> | |||
Regular Zymo cleanup<br> | |||
Ligation<br> | |||
Mistakes on measuring the volume of the insert<br> | |||
PCR again<br> | |||
<br> | |||
'''Feb 27 2009'''<br> | |||
---- | |||
Digestion (upaG)<br> | |||
Regular zymo cleanup (upaG)<br> | |||
<br> | |||
I finally received the oligos for the second part of the project<br> | |||
mea36&mea37<br> | |||
1.Dilution<br> | |||
2.Regular PCR<br><Br> | |||
'''Mar 2-3 2009'''<br> | |||
---- | |||
Some mistakes occurred on PCR for phoA by GSI<br> | |||
Should re-do it<br><br> | |||
'''Mar 4 2009'''<br> | |||
---- | |||
upaG Long<br> | |||
1.ligation<br> | |||
2.Accidently put it in the thermalcycler<br> | |||
3.Digestion again<br> | |||
4.Run it on the thermalcycler for about 1 hour<br> | |||
phoA<br> | |||
1.Regular PCR again<br> | |||
2.Run on 2k55 since it's under 2k bp (1219bp) | |||
<br> | |||
<br> | |||
'''Mar 9 2009'''<br> | |||
---- | |||
upaG Long<br> | |||
1.Ligation<br> | |||
The cell cocktail was already set up by GSI<br> | |||
Shaking at 37 degree in the incubator<Br> | |||
2.<Br> | |||
<Br> | |||
phoA<br> | |||
1.analytical gel<br> | |||
The result came out bad-the band of my PCR was overlapped with the bands of ladder<br> | |||
Gabe said it would be okay<br> | |||
2.regular zymo cleanup<br> | |||
3.digestion<br> | |||
Latest revision as of 12:05, 9 March 2009
Feb 18 2009
First day of the actual lab.
Lab work on my project.
Template:SBB09 19329
upaG Long
I added 275μL of water in the given tube in which JS001F oligo was and 343μL of water in the JS002R tube.
Then, I followed the instruction below.
Cloning by PCR
Since the size is over 4000K, I put the PCR tube in 8k55.
Feb 23 2009
Second day of the lab
I guess Prof. Chris forgot to give me oligos for Alkaline Phosphatase
I should do it later once i get the oligos
To decide if there's product in the PCR tube I worked on Feb 18
I need to run it on gel
1. Add 5μL of the product and 1μL of loading buffer
2. Insert 6μL in total onto gel
3. Wait for about 10 minutes
4. GSIs take a picture of my analytical gel
6. The picture link for the first product
mine is the 5th one
The size is between 1.5k and 1k; it's about the right size I've expected. Product? Yes.
The picture link for the second product
mine is the 2nd one
The result came out the same as the first one. Product? Yes!
7. Zymo cleanup
Feb 25 2009
Digestion
Regular Zymo cleanup
Ligation
Mistakes on measuring the volume of the insert
PCR again
Feb 27 2009
Digestion (upaG)
Regular zymo cleanup (upaG)
I finally received the oligos for the second part of the project
mea36&mea37
1.Dilution
2.Regular PCR
Mar 2-3 2009
Some mistakes occurred on PCR for phoA by GSI
Should re-do it
Mar 4 2009
upaG Long
1.ligation
2.Accidently put it in the thermalcycler
3.Digestion again
4.Run it on the thermalcycler for about 1 hour
phoA
1.Regular PCR again
2.Run on 2k55 since it's under 2k bp (1219bp)
Mar 9 2009
upaG Long
1.Ligation
The cell cocktail was already set up by GSI
Shaking at 37 degree in the incubator
2.
phoA
1.analytical gel
The result came out bad-the band of my PCR was overlapped with the bands of ladder
Gabe said it would be okay
2.regular zymo cleanup
3.digestion
