10:15a – 5:00p,
remotely, 10:00p-12:00a
Cell staining
- wash out secondary stains.
- no microscope time available.
matlab-storm
- debugging new info-file reader to determine analyzed region
- There is still a problem here. There is only one info file for the movie.
chromewarp issues
- Determining whether ROI is indexed from 0 or 1.
- pixels 0 – 256 corresponds to x-position data
x>0 & x<257? - quadviewsplit in ReadDaxBeta splits 1:256,257:512
- 0-257 and 257-513? 0.5-256, 256 to 512.5
- warp files not copied out of Beta. Should now be fixed.
- finally found real issue with my 2 differently behaving bead-maps: 3D warping in map 2 is not aligning beads correctly. (still not sure why Chromewarps claims small warp error).
- individually analyze STORM movies.
Literature
- Zhang, Y. et al. Chromatin connectivity maps reveal dynamic promoter-enhancer long-range associations. Nature 504, 306–310 (2013).
- http://www.ncbi.nlm.nih.gov/pubmed/23883933.
- “We show that gene promoters engage in contacts between topological domains in a largely tissue-independent manner, whereas enhancers have a more tissue-restricted interaction profile.” – artifact of local chromatin accessibility of promoters?
- Very interesting: “Different from what is observed in somatic cells, in embryonic stem cells we find that transcriptionally inactive regions form low numbers of specific long-range contacts (Fig. 1a, b). This is not due to their inability to reach over large distances, but instead to a more random organization of their long-range captures”
- Why wasn’t this noticed / discussed in the previous studies? — (that inactive chromatin makes more robust contacts in mature cells compared to ES cells) “The Hi-C data confirm on a global scale that inactive and active chromatin differ in their propensity to form specific long-range contacts in ES cells”
- iPS cells, “A reactivated gene (Nanog) gains contacts during iPS cell reprogramming, whereas a resilenced gene (Ptprz1) loses all but two contacts (Supplementary Fig. 10)” – really suggesting more contacts for active genes. Doesn’t seem to be in qualitative agreement with HiC maps — much more extended domains seen for silent (Blue/Green/Black) regions.
- gene sequence of Nanog forms specific contacts with other pluripotency factor gene loci in an ES-cell specific manner. (Also true for sox2 enhancer)
- DNA FISH done with 200+ kb BACs (Nanog BAC is RP23-117I23, 221.8 kb). Average insert size in library is 197 kb. See here
- Typical 4C maps are on the scale of 40 – 150 Mb in this paper.