I am looking for a light-triggered base generator or something that increases the pH upon light irradiation(380-405nm), that is commercially available.
Recently there have been several studies that show, using a truncated sgRNA with a wild type CAS9 can deter the nuclease activity and can potentially be used for epigenetic regulation.
Kiani et al.(2015) claimed that 'that the lack of DNA cleavage with 16-nt gRNA was due not to a lack of DNA binding but to an inability of Cas9 to cleave the target substrate after binding.'
I wish to know how and why does the size of sgRNA affects the nuclease activity.
I want to know what is the best protocol for staining PBMCs on glass slides with wright giemsa stain. Does the basic smearing with another slide cause lysis of cells? Also, what is the best technique to fix the cells?
I recently tried extracting DNA from hair.. i took samples from 5 different individuals in different eppendorfs. I used B-Mercaptoethanol(instead of DTT(X10 molarity)) and proteinaseK for dissolution of keratin... i used the protocol given in the link..
i followed the protocol properly and i got decent amount of white pellet after NaOAc precipitation and after washing with ethanol.
when i loaded it for gel electrophoresis i got no bands.. nothing at all.. no protein impurity... nothing in all 5 samples..
and side by side i also extracted Dna from saliva.. i got the same result.. no bands..
what am i doing wrong?