Arichard:Notebook/2013/November
November, 2013[edit]
- Following CSHL Nat Prot for CNV analysis, generated 50 base reads, 1 from each position of mm10.
- Mapped back to mm10 using bowtie.
- Files are on tscc under /oasis/tscc/scratch/arichard/mouseCNV
11/22/2013[edit]
Motivation: We have condensed (metaphase arrested) PGP1 chromosomes suspended at 4 degC in PBS (as far as I know, only PBS). They have been in storage for at least a year. We attempted a haplotyping experiment several months ago and got poor results due to low mapping. This may have been related to the low mapping issue we have had with nuclei, but we also suspect that the chromosomes may have degraded and/or relaxed during storage.
Objective: Stain PGP1 chromosomes with SYBR Green and determine whether they are visible as puncta at 40X objective.
This is H2O with 1:9 SYBR Green at 40X
File:20131122 pgp1 1to9 sybGreen 40xMag neg.tif
This is PGP1 chromosomes with 1:9 SYBR Green at 40X
File:20131122 pgp1 1to9 sybGreen 40xMag.tif
This is H2O with 1:14 SYBR Green at 40X
File:20131122 pgp1 1to14 sybGreen 40xMag neg.tif
This is PGP1 chromosomes with 1:14 SYBR Green at 40X
File:20131122 pgp1 1to14 sybGreen 40xMag.tif
There is no difference between negative controls and experimental. We do not have a positive control for intact, condensed chromosomes. I could use bacteria. The takeaway from this test, however, is that there are no intact chromosomes visible with SYBR Green at 40X objective.