tag:blogger.com,1999:blog-3017697080484068141.post7391142410096370928..comments2023-10-08T04:37:34.635-07:00Comments on No DNA Control: As easy as that? Nahh...Changhttp://www.blogger.com/profile/12291718994939895064noreply@blogger.comBlogger2125tag:blogger.com,1999:blog-3017697080484068141.post-42559715166329820632009-05-31T16:18:23.063-07:002009-05-31T16:18:23.063-07:00I'd forgotten that about lambda...
We'd...I'd forgotten that about lambda... <br /><br />We'd ideally use several sizes of uptake input DNA. I'd say 500 bp is the size we should start with. But a large size, say 3 kb would be pretty useful too, but would require us to do some additional downstream processing before we could sequence. Apparently, Solexa is happy with fragments 200 bp - 1 kb, but the larger the fragment, the bigger the cluster, so the fewer total reads per lane you get. Apparently the optimum is somewhere in the few hundred bases range. Or so I hear.<br /><br />We really do need everything to be in a pretty tight size distribution by the end, as it will apparently make the cluster synthesis more uniform.Changhttps://www.blogger.com/profile/12291718994939895064noreply@blogger.comtag:blogger.com,1999:blog-3017697080484068141.post-12132322325072414742009-05-31T07:56:26.627-07:002009-05-31T07:56:26.627-07:00Nice!
Our synthetic uptake fragments are only abo...Nice!<br /><br />Our synthetic uptake fragments are only about 200 bp. The fragments we'd use in the experiments could be bigger, right?<br /><br />The MAP7 DNA was prepared with no attention to protecting it from shearing - it's been vigorously pipetted and probably vortexed. For your experiments the chromosomal DNA would be much bigger because you'd treat the prep very gently.<br /><br />p.s. If you didn't heat the lambda DNA to 65C before using it, the largest fragment is actually two, 23 kb and 28kb, the latter formed by sticky-end annealing of the 23 and 4.4 kb fragments.Rosie Redfieldhttps://www.blogger.com/profile/06807912674127645263noreply@blogger.com