Hey guys, I am new to this community, and was wondering if you could help me out with something
Recently received a BAC clone from CHORI and I am trying to "prove" that they sent us the appropriate BAC. Using solution based mini prep (not a kit) I have extracted the BAC, on spec it shows ~700ng/ul with a 260/280 of ~1.9. I have 2 BAC clones of different sizes, both are rather large (one is >50kb) - although I would need to pull up the exact numbers if that is important for you to help. Using GCK I have predicted the restriction digest for ECOR1 which shows a few small bp bands (one ~250 and one ~500bp). I have been running on a 1% agarose gel and my problem has been that I do not see any bands. The DNA ladders look as expected, but in the wells containing the digested BAC clone I don't see any bands. Looking back at my notes, I have been loading too little DNA for the 500bp bands to light up, but I can't figure out why the larger bands ~5kb (or larger) aren't showing up at all? I would expect there to be at least 20ng of the 5kb bands present in the well, and I am using ethidium bromide to stain.
I have also run an undigested sample which did not show any bands either.
I feel confident that there is DNA present in the well - what could be going on here?
Thank you for your time and consideration!