Haynes Lab:Notebook/CRISPR Editing/2015/03/03: Difference between revisions

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|style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;"> Project name</span>
|style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;"> Project name</span>
|style="background-color: #F2F2F2" align="center"|<html><img src="/images/9/94/Report.png" border="0" /></html> [[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|<html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>[[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]]<html>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;</html>}}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]]<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html>}}
|style="background-color: #F2F2F2" align="center"|[[File:Report.png|frameless|link={{#sub:{{FULLPAGENAME}}|0|-11}}]][[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|[[File:Resultset_previous.png|frameless|link={{#lnpreventry:{{FULLPAGENAME}}}}]][[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]]&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;}}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]][[File:Resultset_next.png|frameless|link={{#lnnextentry:{{FULLPAGENAME}}}}]]}}
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==02//2015==
==03/03/2015==
 
Checked concentrations on minipreps from last year of Lonza pMax GFP from Krishna. very low so JW transformed the original along with pMax GFP 6.2 and KAH60.
 
{| {{table}}
 
| align="center" style="background:#f0f0f0;"|'''Sample Read#'''
| align="center" style="background:#f0f0f0;"|'''260'''
| align="center" style="background:#f0f0f0;"|'''280'''
| align="center" style="background:#f0f0f0;"|'''260/280'''
| align="center" style="background:#f0f0f0;"|'''ng/µL'''
|-
| pMax GFP||0.026||0.015||1.736||26.294
|-
| pMax GFP||0.023||0.013||1.766||23.175
|-
| pMax GFP||0.031||0.019||1.639||30.851
|}
<br><br>
Ran PCR products from P151/158 on gel. Only single product for each band.
<br><br>
Sequencing:<br>Need to purify. Will purify 25ul and elute into 35 ul.<br>
Purification results:
{| {{table}}
| align="center" style="background:#f0f0f0;"|'''Cell type'''
| align="center" style="background:#f0f0f0;"|'''treatment'''
| align="center" style="background:#f0f0f0;"|'''rep'''
| align="center" style="background:#f0f0f0;"|'''260'''
| align="center" style="background:#f0f0f0;"|'''280'''
| align="center" style="background:#f0f0f0;"|'''260/280'''
| align="center" style="background:#f0f0f0;"|'''ng/µL'''
|-
| Luc14||Untreated||1||0.035||0.017||2.006||35.007
|-
| Luc14||g046||1||0.032||0.016||2.02||31.691
|-
| Luc14||Untreated||2||0.045||0.022||2.034||44.797
|-
| Luc14||Untreated||3||0.046||0.024||1.938||46.277
|-
| Gal4-EED +dox||Untreated||1||0.04||0.019||2.099||40.212
|-
| Gal4-EED +dox||Untreated||2||0.043||0.023||1.885||42.773
|-
| Gal4-EED +dox||Untreated||3||0.044||0.023||1.93||43.675
|-
| Luc14||g046||2||0.03||0.015||2.086||30.259
|-
| Luc14||g046||3||0.031||0.015||2.113||31.368
|-
| Gal4-EED +dox||g046||1||0.035||0.017||2.042||35.122
|-
| Gal4-EED +dox||g046||2||0.039||0.019||2.05||38.501
|-
| Gal4-EED +dox||g046||3||0.041||0.022||1.897||41.336
|-
| Luc14||Untreated||1||0.032||0.016||1.949||31.937
|-
| Luc14||g046||1||0.03||0.015||1.94||29.645
|}
<br>
Sent for sequencing at biodesign with P176
{| {{table}}
| align="center" style="background:#f0f0f0;"|'''Label'''
| align="center" style="background:#f0f0f0;"|'''Cell type'''
| align="center" style="background:#f0f0f0;"|'''treatment'''
| align="center" style="background:#f0f0f0;"|'''rep'''
| align="center" style="background:#f0f0f0;"|'''PCR primer'''
|-
| 1||Luc14||Untreated||1||P176
|-
| 2||Luc14||Untreated||2||P176
|-
| 3||Luc14||Untreated||3||P176
|-
| 4||Gal4-EED+dox||Untreated||1||P176
|-
| 5||Gal4-EED+dox||Untreated||2||P176
|-
| 6||Gal4-EED+dox||Untreated||3||P176
|-
| 7||Luc14||g034||1||P176
|-
| 8||Luc14||g034||2||P176
|-
| 9||Luc14||g034||3||P176
|-
| 10||Gal4-EED+dox||g034||1||P176
|-
| 11||Gal4-EED+dox||g034||2||P176
|-
| 12||Gal4-EED+dox||g034||3||P176
|-
| 13||Luc14||g046||1||P176
|-
| 14||Luc14||g046||2||P176
|-
| 15||Luc14||g046||3||P176
|-
| 16||Gal4-EED+dox||g046||1||P176
|-
| 17||Gal4-EED+dox||g046||2||P176
|-
| 18||Gal4-EED+dox||g046||3||P176
|}
<br><br>
Ran PCR with P151/158 on g023, g045, g048 PCR with P197/198 diluted 1:1000 from yesterday. Ran on gel.
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|}
|}


__NOTOC__
__NOTOC__

Latest revision as of 00:48, 27 September 2017

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03/03/2015

Checked concentrations on minipreps from last year of Lonza pMax GFP from Krishna. very low so JW transformed the original along with pMax GFP 6.2 and KAH60.

Sample Read# 260 280 260/280 ng/µL
pMax GFP 0.026 0.015 1.736 26.294
pMax GFP 0.023 0.013 1.766 23.175
pMax GFP 0.031 0.019 1.639 30.851



Ran PCR products from P151/158 on gel. Only single product for each band.

Sequencing:
Need to purify. Will purify 25ul and elute into 35 ul.
Purification results:

Cell type treatment rep 260 280 260/280 ng/µL
Luc14 Untreated 1 0.035 0.017 2.006 35.007
Luc14 g046 1 0.032 0.016 2.02 31.691
Luc14 Untreated 2 0.045 0.022 2.034 44.797
Luc14 Untreated 3 0.046 0.024 1.938 46.277
Gal4-EED +dox Untreated 1 0.04 0.019 2.099 40.212
Gal4-EED +dox Untreated 2 0.043 0.023 1.885 42.773
Gal4-EED +dox Untreated 3 0.044 0.023 1.93 43.675
Luc14 g046 2 0.03 0.015 2.086 30.259
Luc14 g046 3 0.031 0.015 2.113 31.368
Gal4-EED +dox g046 1 0.035 0.017 2.042 35.122
Gal4-EED +dox g046 2 0.039 0.019 2.05 38.501
Gal4-EED +dox g046 3 0.041 0.022 1.897 41.336
Luc14 Untreated 1 0.032 0.016 1.949 31.937
Luc14 g046 1 0.03 0.015 1.94 29.645


Sent for sequencing at biodesign with P176

Label Cell type treatment rep PCR primer
1 Luc14 Untreated 1 P176
2 Luc14 Untreated 2 P176
3 Luc14 Untreated 3 P176
4 Gal4-EED+dox Untreated 1 P176
5 Gal4-EED+dox Untreated 2 P176
6 Gal4-EED+dox Untreated 3 P176
7 Luc14 g034 1 P176
8 Luc14 g034 2 P176
9 Luc14 g034 3 P176
10 Gal4-EED+dox g034 1 P176
11 Gal4-EED+dox g034 2 P176
12 Gal4-EED+dox g034 3 P176
13 Luc14 g046 1 P176
14 Luc14 g046 2 P176
15 Luc14 g046 3 P176
16 Gal4-EED+dox g046 1 P176
17 Gal4-EED+dox g046 2 P176
18 Gal4-EED+dox g046 3 P176



Ran PCR with P151/158 on g023, g045, g048 PCR with P197/198 diluted 1:1000 from yesterday. Ran on gel.


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