You said that you cleave the larger DNA fragments into smaller gene fragments... but if the functions are not known, how do you known when you're cutting a gene itself out and not halfway though a gene? Do you look for start/ stop codons first or something? And how do you separate the genes into different beakers, is it based on differential centrifugation...? I'm a little confused!
yes im confused as well. im assuming you can take the dna that has been run through the gel and then clone it and put them in beaker? i might be wrong, ive never run the gel electrophoresis before
you have to carry out partial dna digestion to improve your chances of having atleast one intact copy of each gene because indeed restriction enzymes don't respect gene boundaries. Later on you can look if the gene of interest is present by using a complementary strand.
Does the restriction enzyme cut precisely one gene at a time or does the fragment sometimes include more (or less) than one gene? How does the restriction recognize where to cut?
Isn't it an audacious claim to make when you say that the restriction sites are present exactly at the intersection of 2 genes? I believe the restriction site locations will be extremely impossible to predict, as we never know where a palindromic sequence is supposed to be.
As far as I know, scientists use many different restriction enzymes on different copies of one genome to be sure to have the maximum representation (and to include the sequence they're seeking)
Do we still (now) need a phage vector to replicate the gene fragments? Why can't we do a PCR to amplify the gene fragments in large number followed by insertion into e.coli and screen for our gene of interest?
can you tell me how the probe is going to get hybridised with the dna of interest?and how we are going to seperate that hybridised dna for further process?
How many recombinants has to be screened with 90 percent probbality of getting a gene if the size of genome is 8000kb and library was constructed in puc vector Please answer
