A+G Maxam Gilbert ladder for predicting transcription start site - (Aug/31/2008 )
I have to find out transcription start site of a prokaryotic gene. I have seen a paper in which auther, after doing S1 nuclease assay, has run the undigested hyberdigestion products on a gel while using A+G Maxam Gilbert ladder a marker. Only with this, How can be exact transcription start site predicted?
Well, you (presumably) already know the sequence. The A+G lane simply gives you a marker lane which lets you calibrate the (known) sequence against the location of the transcription start site. It would not work unless you already knew the DNA sequence some other way (presumbably with Sanger ddNTP sequencing).
Thanks a lot for guiding. Yet I am still not clear about This ladder.
Briefly, what I have plan is that I have to hybridize RNA with some gene specific labelled probe, subject it to S1 nuclease assay and then run it on gel while using this ladder as marker. What I understand is that by comparing the position of the band against this ladder will tell the size of the hybrid. Thus we can determine the transcription start site.
Secondly I do not know whether this ladder is available commercially and if yes then from which company. Does Sigma provides this. As usually my department buys chemicals and reagents from Sigma.
Please do guide me in some detail as early as possible so that I can place my order with other people within a few days.