Glasgow/Wetlab/Week3

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[[Glasgow|Glasgow Main Page]] | [[Glasgow/Wetlab|Back To Wetlab Log]]
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!align=center|[[Image:Uog.jpg]] ||  [[Glasgow|<font face=georgia color=#3366CC size=4>Back To <br> Glasgow's <br> Main Page</font>]] || [[Glasgow/Wetlab|<font face=georgia color=#3366CC size=4>Back To <br> Glasgow's <br> Wetlab Log</font>]] ||  [[Glasgow/Modeling|<font face=georgia color=#3366CC size=4>Go To <br> Glasgow's <br>Drylab Log</font>]]
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{|cellspacing="6px" cellpadding="16" border="0" width="100%"
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|[https://2007.igem.org/Glasgow/Wetlab/Protocols <font face=georgia color=#3366CC size=5><b>Protocols</b></font>]
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|[https://2007.igem.org/Glasgow/Wetlab/References <font face=georgia color=#3366CC size=5><b>References</b></font>]
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|[https://2007.igem.org/Glasgow/Wetlab/Resources <font face=georgia color=#3366CC size=5><b>Resources</b></font>]
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|[https://2007.igem.org/Glasgow/Wetlab/Orders <font face=georgia color=#3366CC size=5><b>Orders</b></font>]
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|[https://2007.igem.org/Glasgow/Wetlab/Biobricks <font face=georgia color=#3366CC size=5><b>Biobricks<br>Used</b></font>]
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|[https://2007.igem.org/Glasgow/Wetlab/Gels <font face=georgia color=#3366CC size=5><b>Gels</b></font>]
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|BBa_I52001
|BBa_I52001
| p5B4K5
| p5B4K5
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|High copy death gene
|AvaI, PvuI
|AvaI, PvuI
|920bp, 1470bp, 2120bp
|920bp, 1470bp, 2120bp
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#Last night we diluted primers to 100 pmol/ul by adding the amount of dH2O given on the order sheet, and made working solutions to 10 pmol/ul (2 x 200 ul each - 20 ul stock and 180 ul dH2O).
#Last night we diluted primers to 100 pmol/ul by adding the amount of dH2O given on the order sheet, and made working solutions to 10 pmol/ul (2 x 200 ul each - 20 ul stock and 180 ul dH2O).
#[[User:MaijaP|Maija]] ran a gel of the restriction digests to check the sizes of the biobricks.  These were the maps we made yesterday.
#[[User:MaijaP|Maija]] ran a gel of the restriction digests to check the sizes of the biobricks.  These were the maps we made yesterday.
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#[[User:Mojs|Maia]] is extracting DNA from 3 samples of ****, which were plated yesterday and grown in liquid media, using MoBio PowersoilTM DNA purification kit (see [[Glasgow/Wetlab/Protocols#Protocol 8: MoBio PowersoilTM DNA Purification Kit|Protocol 8]]); only change was to shorten "shaking" time from 10 to 2 minutes.
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#[[User:Mojs|Maia]] is extracting DNA from 3 samples of Pseudomonas Aeruginosa, which were plated yesterday and grown in liquid media, using MoBio PowersoilTM DNA purification kit (see [[Glasgow/Wetlab/Protocols#Protocol 8: MoBio PowersoilTM DNA Purification Kit|Protocol 8]]); only change was to shorten "shaking" time from 10 to 2 minutes.
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#PCR trial run using Reddymix and Touch 2 done on:
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#PCR trial run using Reddymix and Touch 2 done (see [[Glasgow/Wetlab/Protocols#Protocol 9: PCR|Protocol 9]] for PCR).
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#*pGLTUR: XylR_prefix / XylR_suffix, Pr_prefix / Pr_suffix, Pr_prefix / XylR_suffix, Pu_prefix_EM / Pu_suffix_EM.
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{| border="1" cellspacing="0" cellpadding="5" align="center"
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#*pQF52 plasmid: DntR_prefix / DntR_suffix
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!Template
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#*DmpR WT/24: DntR_prefix / DntR_suffix
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!Primers
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|-
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|pGLTUR  
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|
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<br>XylR_prefix + XylR_suffix
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<br>Pr_prefix + Pr_suffix  
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<br>Pr_prefix + XylR_suffix
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<br>Pu_prefix_EM + Pu_suffix_EM
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|-
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|pQF52  
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|DntR_prefix + DntR_suffix
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|-
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|DmpR WT/24  
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|DntR_prefix + DntR_suffix
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|}
#*Biobricks: [http://partsregistry.org/Part:BBa_R0062 R0062], [http://partsregistry.org/Part:BBa_E0040 E0040], [http://partsregistry.org/Part:BBa_J04500 J04500], [http://partsregistry.org/Part:BBa_J23119 J23119], [http://partsregistry.org/Part:BBa_p1010 p1010], [http://partsregistry.org/Part:BBa_I52001 I52001].
#*Biobricks: [http://partsregistry.org/Part:BBa_R0062 R0062], [http://partsregistry.org/Part:BBa_E0040 E0040], [http://partsregistry.org/Part:BBa_J04500 J04500], [http://partsregistry.org/Part:BBa_J23119 J23119], [http://partsregistry.org/Part:BBa_p1010 p1010], [http://partsregistry.org/Part:BBa_I52001 I52001].
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#*See [[Glasgow/Wetlab/Protocols#Protocol 9: PCR|Protocol 9]] for PCR.
 
=== Thursday 19th July 2007 ===
=== Thursday 19th July 2007 ===
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#[[User:Mojs|Maia]] redid PCR on pGLTUR and pQF52 using Reddymix and a gradient program (See [[Glasgow/Wetlab/Protocols#Protocol 9: PCR|Protocol 9]]) to minimise the non-specific amplification evident in previous gels. New DntR suffix primer has arrived. Used the following primer sets.
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#[[User:Mojs|Maia]] redid PCR on pGLTUR and pQF52 using Reddymix and a gradient program (see [[Glasgow/Wetlab/Protocols#Protocol 9: PCR|Protocol 9]]) to minimise the non-specific amplification evident in previous gels. New DntR suffix primer has arrived. Used the following primer sets.
#*XylR prefix and XylR suffix
#*XylR prefix and XylR suffix
#*Pr prefix and Pr suffix
#*Pr prefix and Pr suffix
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=== Friday 20th July 2007 ===
=== Friday 20th July 2007 ===
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<br>
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![[Glasgow/Wetlab/Week2|<font face=georgia color=#3366CC size=4>Previous  <br>  Week</font>]] || [[Glasgow/Wetlab/Week4|<font face=georgia color=#3366CC size=4>Next <br>  Week</font>]]
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{| valign=top cellpadding=3
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|-
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!align=center|[[Image:Uog.jpg]] ||  [[Glasgow|<font face=georgia color=#3366CC size=4>Back To <br> Glasgow's <br> Main Page</font>]] || [[Glasgow/Wetlab|<font face=georgia color=#3366CC size=4>Back To <br> Glasgow's <br> Wetlab Log</font>]]
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Latest revision as of 17:08, 22 October 2007

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Glasgow's
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Wetlab Log 		Go To
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Drylab Log
Protocols References Resources Orders Biobricks
Used 		Gels

Contents

Week 3

Tuesday 17th July 2007

  1. Restriction Digests of transformed BioBricks (see Protocol 7):
Label BioBrick Plasmid Description Enzymes Expected Sizes
3/19A BBa_J23119 pSB1A2 strong constitutive promoter NheI, PvuI 680bp, 1430bp
1/9G BBa_R0062 pBB1A2 HSL and luxR EcoRI, PvuI 1460bp, 660bp
1/16P BBa_J04500 pSB1AK3 IPTG inducer and RBS PvuII, PvuI 2250bp, 1030bp, 730bp
4/11C BBa_p1010 pSB3K3 death gene BamHI, XhoI 190bp, 2390bp,840bp
1/5H BBa_E0040 pSB1A2 GFP no promoter Hime II, PvuI 1630bp, 1170bp
4/6D BBa_I52001 p5B4K5 High copy death gene AvaI, PvuI 920bp, 1470bp, 2120bp

Wednesday 18th July 2007

  1. Last night we diluted primers to 100 pmol/ul by adding the amount of dH2O given on the order sheet, and made working solutions to 10 pmol/ul (2 x 200 ul each - 20 ul stock and 180 ul dH2O).
  2. Maija ran a gel of the restriction digests to check the sizes of the biobricks. These were the maps we made yesterday.
  3. Maia is extracting DNA from 3 samples of Pseudomonas Aeruginosa, which were plated yesterday and grown in liquid media, using MoBio PowersoilTM DNA purification kit (see Protocol 8); only change was to shorten "shaking" time from 10 to 2 minutes.
  4. PCR trial run using Reddymix and Touch 2 done (see Protocol 9 for PCR).
Template Primers
pGLTUR


XylR_prefix + XylR_suffix
Pr_prefix + Pr_suffix
Pr_prefix + XylR_suffix
Pu_prefix_EM + Pu_suffix_EM

pQF52 DntR_prefix + DntR_suffix
DmpR WT/24 DntR_prefix + DntR_suffix

Thursday 19th July 2007

  1. Maia redid PCR on pGLTUR and pQF52 using Reddymix and a gradient program (see Protocol 9) to minimise the non-specific amplification evident in previous gels. New DntR suffix primer has arrived. Used the following primer sets.
    • XylR prefix and XylR suffix
    • Pr prefix and Pr suffix
    • Pr prefix and XylR suffix
    • Pu prefix and Pu suffix
    • DntR prefix and DntR suffix 2

Friday 20th July 2007


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