Hi,everyone
I am quite fresh in protein expression. Now, I expressed a small peptide(0.8KD) in Pichia pastoris with the vector pPICZ alpha. It is not the fusion expression with his-tag. But the purification is difficult and I cannot detect the peptide in the fermentation culture. I do not know whether the expression or the purification is some wrong.
Thanks in advance for help.
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#2
zhgljj1998
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Posted 28 February 2005 - 08:29 AM
design another peptide with Tyr, other AA are same. then you got idea to get concentration of your small peptide.
#3
sharath
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Posted 28 February 2005 - 10:59 PM
If you have a sample of the peptide you are trying to express then you can run your extract along with the authentic in SDS PAGE and check for its presence.
Otherwsie, if you peptide has some bioassay or activity, then you can use you extract to check for it. You need a means of monitoring your peptide during purification. How do you plan to do?
Begin your purifcation only after confirming the presence of your peptide in the initial extract.
Write about the purification protocol that you employ. Lets see if there is any source of loss in it.
good luck
Otherwsie, if you peptide has some bioassay or activity, then you can use you extract to check for it. You need a means of monitoring your peptide during purification. How do you plan to do?
Begin your purifcation only after confirming the presence of your peptide in the initial extract.
Write about the purification protocol that you employ. Lets see if there is any source of loss in it.
good luck
Edited by sharath, 01 March 2005 - 10:11 AM.
Sharath B.
