Matt:LabNotes/2013-6-4: Difference between revisions

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===Probe Capture===
===Probe Capture===
*Use the ~37.4 pmole of probes from [http://genome-tech.ucsd.edu/LabNotes/index.php/Matt:LabNotes/2013-5-22] and [http://genome-tech.ucsd.edu/LabNotes/index.php/Matt:LabNotes/2013-5-29] to produce maximum number of circularized probes with U's incorporated for Ho Suk's experiments
*From [http://genome-tech.ucsd.edu/LabNotes/index.php/Matt:LabNotes/2013-5-22] and [http://genome-tech.ucsd.edu/LabNotes/index.php/Matt:LabNotes/2013-5-29]: ~37.4 pmol in 140ul
*Use gDNA as template/target
*Use gDNA as template/target
*Use a probe:target ratio of 100:1
*Use a probe:target ratio of 100:1


====100:1 Probe:Target Calculations====
*Hosuk said there are about 3,000 cells per dish, I will round up to 10,000 so there are extra oligos for multiple experiments


10^4 cells x 10^4 rolonies/cell = 10^8 rolonies in dish <br>
10^8 rolonies / 4,000 genes = 2.5 x 10^4 rolonies/gene (targets) <br>
100 x 2.5 x 10^4 = 2.5 x 10^6 probe sets => 4.151 x 10^-18 mol of probe sets


56 ug of gDNA
====100:1 Probe:Target Calculations====
140 ul of Probes
{| {{table}}
| Probe set size||12964||probes
|-
| gDNA template||300||ng
|-
| gDNA MW||1.95E+12||g/mol (3x10^9 bp x 650 Da/bp + 157.9 Da)
|-
| template||1.5385E-19||mol
|-
| probe sets (100:1)||1.5385E-17||mol
|-
| probe MW (12,964 probes, 193nt)||814190000||g/mol (12,964 probes x (193 nt/probe x 325 Da/nt + 79 Da))
|-
| amount probe required||1.25263E-08||g
|-
| ||12.5||ng
|-
|
|}

Revision as of 00:36, 5 June 2013

Continued from: http://genome-tech.ucsd.edu/LabNotes/index.php/Matt:LabNotes/2013-5-22

Probe Capture

  • From [1] and [2]: ~37.4 pmol in 140ul
  • Use gDNA as template/target
  • Use a probe:target ratio of 100:1
  • Hosuk said there are about 3,000 cells per dish, I will round up to 10,000 so there are extra oligos for multiple experiments

10^4 cells x 10^4 rolonies/cell = 10^8 rolonies in dish
10^8 rolonies / 4,000 genes = 2.5 x 10^4 rolonies/gene (targets)
100 x 2.5 x 10^4 = 2.5 x 10^6 probe sets => 4.151 x 10^-18 mol of probe sets

100:1 Probe:Target Calculations

Probe set size 12964 probes
gDNA template 300 ng
gDNA MW 1.95E+12 g/mol (3x10^9 bp x 650 Da/bp + 157.9 Da)
template 1.5385E-19 mol
probe sets (100:1) 1.5385E-17 mol
probe MW (12,964 probes, 193nt) 814190000 g/mol (12,964 probes x (193 nt/probe x 325 Da/nt + 79 Da))
amount probe required 1.25263E-08 g
12.5 ng