Daniel:Notebook/HiResChrPaint/2014-7-18: Difference between revisions

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=Dystrophin Probes=
=Dystrophin Probes=


[[Daniel:Notebook/HiResChrPaint/2014-7-18|Back to Calendar]]
[[Daniel:Notebook/HiResChrPaint|Back to Calendar]]


The dystrophin probes I designed [[Daniel:Notebook/HiResChrPaint/2014-7-10|last week]] arrived from IDT.  Attached are their specs:
The dystrophin probes I designed [[Daniel:Notebook/HiResChrPaint/2014-7-10|last week]] arrived from IDT.  Attached are their specs:
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==Oligo Pool Synthesis==
==Oligo Pool Synthesis==


#Resuspend each oligo to make them all 100 uM (See Plate Specs for values)
#Resuspend each oligo to make them all 1 mM (See Plate Specs for values)
#Combine 20 uL from each oligo, makes 2 ug/uL total pool
#Combine 2 uL from each oligo  
#Add 640 uL nfH2O, makes 1 ug/uL total pool
##Makes 12.5 ug/uL total pool
##Total volume will be 64 uL
#Add 742.4 uL nfH2O, dilutes to 1 ug/uL total pool
##Create dilution series and measure in nanodrop
 
===Nanodrop Results===
 
{| class="wikitable" <hiddentext>generated with [[:de:Wikipedia:Helferlein/VBA-Macro for EXCEL tableconversion]] V1.8</hiddentext>
|- style="font-size:12pt;font-weight:bold" align="center" valign="bottom"
| width="78" height="15" | Dilution (X:1)
| width="65" | ng/uL
| width="65" | proj. ug/uL
 
|- style="font-size:12pt" align="center" valign="bottom"
| align="center" height="15" | 5
| align="center" | 79.6
| align="center" | 0.40
 
|- style="font-size:12pt" align="center" valign="bottom"
| align="center" height="15" | 10
| align="center" | 38.2
| align="center" | 0.38
 
|- style="font-size:12pt" align="center" valign="bottom"
| align="center" height="15" | 20
| align="center" | 19.3
| align="center" | 0.39
 
|- style="font-size:12pt" align="center" valign="bottom"
| align="center" height="15" | 50
| align="center" | 7.4
| align="center" | 0.37
 
|- style="font-size:12pt" align="center" valign="bottom"
| align="center" height="15" | 100
| align="center" | 3.7
| align="center" | 0.37
 
|- style="font-size:12pt" align="center" valign="bottom"
|style="font-weight:bold" height="15" | Average
| align="center" | &nbsp;
|style="font-weight:bold" align="center" | 0.38
 
|}
 
So I guess my calculations were a bit off, but 0.4 ug/uL is pretty good for FISH, especially considering I need 30 pmol per experiment (0.4 ug/uL ~60 pmol/uL).
 
[[Category:HRCP]] [[Category:20140718]]

Latest revision as of 21:19, 19 July 2014

Dystrophin Probes[edit]

Back to Calendar

The dystrophin probes I designed last week arrived from IDT. Attached are their specs:

The oligos arrived as dry oligos in a plate. The calculations I made are in the following text file:

Oligo Pool Synthesis[edit]

  1. Resuspend each oligo to make them all 1 mM (See Plate Specs for values)
  2. Combine 2 uL from each oligo
    1. Makes 12.5 ug/uL total pool
    2. Total volume will be 64 uL
  3. Add 742.4 uL nfH2O, dilutes to 1 ug/uL total pool
    1. Create dilution series and measure in nanodrop

Nanodrop Results[edit]

Dilution (X:1) ng/uL proj. ug/uL
5 79.6 0.40
10 38.2 0.38
20 19.3 0.39
50 7.4 0.37
100 3.7 0.37
Average   0.38

So I guess my calculations were a bit off, but 0.4 ug/uL is pretty good for FISH, especially considering I need 30 pmol per experiment (0.4 ug/uL ~60 pmol/uL).