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Old 12-13-2019, 03:25 AM   #2
Shahrukh
Junior Member
 
Location: Russia

Join Date: Oct 2019
Posts: 2
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So this week, I tried sonication with Sonics Vibra cell 500/750. I want to share my experience and hopefully get input from you. The details are as follows:
- ~3m HEK293T cells per sample
- 500ul RIPA buffer per sample
- 1/8" tapered microtip from Sonics
The sonication conditions I used on four technical replicates are:
A = 3 sec on, 30 sec off at 30%amp for 10 min
B = 3 sec on, 30 sec off at 30%amp for 20 min
C = 3 sec on, 30 sec off at 20%amp for 20 min
D = 3 sec on, 30 sec off at 25%amp for 20 min

All of them had foaming except for sample C, which was sonicated at 20% amplitude. DNA was extracted by using a local Russian kit. I got the following amounts for DNA fragments without being IP'd.
A = 0.33ug
B = 0.32ug
C = 0.38ug
D = 0.26ug
Attached is the gel (sorry our gel doc broken camera is broken). I believe I can play a bit more with the conditions described in C by decreasing the total time to 15 and 10min. Please let me know what'd be suitable.
Also, what's with the low conc.? I didn't even IP'd them. Is it due to less number of cells? Any suggestion to increase the DNA amount.
Attached Images
File Type: jpeg Sonication.jpeg (23.8 KB, 7 views)

Last edited by Shahrukh; 12-13-2019 at 03:36 AM.
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