Glasgow/Wetlab/Week1

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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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----
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{|cellspacing="6px" cellpadding="16" border="0" width="100%"
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|- align=center
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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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|}
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== Week 1 ==
== Week 1 ==
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=== Tuesday 3 July 2007===
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=== Tuesday 3rd July 2007===
#[[User:MaijaP|Maija]] and [[User:Christinemerrick|Christine]] prepared LB broth and LB agar with [[Glasgow/Wetlab/Protocols# Protocol 1: LB Broth and Agar|Protocol 1]].  Made 4 batches of plates, 2 with kanamycin and 2 with carbenicillin.
#[[User:MaijaP|Maija]] and [[User:Christinemerrick|Christine]] prepared LB broth and LB agar with [[Glasgow/Wetlab/Protocols# Protocol 1: LB Broth and Agar|Protocol 1]].  Made 4 batches of plates, 2 with kanamycin and 2 with carbenicillin.
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=== Wednesday 4 July 2007===
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=== Wednesday 4th July 2007===
#All wetlab researched BioBricks.
#All wetlab researched BioBricks.
#Reporter constructs and mini-Tn5 stocks looked out.
#Reporter constructs and mini-Tn5 stocks looked out.
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#*pUJ8 (Carb. Plate)
#*pUJ8 (Carb. Plate)
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=== Thursday 5 July 2007 ===
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=== Thursday 5th July 2007 ===
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#BioBricks – [[User:MaijaP|Maija]] and [[User:scott.w.ramsay|Scott]] transformed using [[Glasgow/Wetlab/Protocols#Protocol 2: Transforming Biobricks | Protocol 2]].
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<ol>
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#*BBa_p1010 (DB3.1) (death gene plasmid) Plate 4: 7A – p5B1A10, 11E – p5B1A10 and 11C – p5B3K3
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<li>BioBricks – [[User:MaijaP|Maija]] and [[User:scott.w.ramsay|Scott]] transformed using [[Glasgow/Wetlab/Protocols#Protocol 2: Transforming Biobricks | Protocol 2]].
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#*BBa_IS2001 (Top10) (high copy number plasmid) Plate 4: 5I – p5B4A5, 5D – p5B4K5, 6B – p5B3K5, 6D – p5B4K5 and 6E – p5B4A5 (Also 5K, 5M, 4J, 4L, 4N, 4P)
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#*BBa_J23119 (Top 10) (strong constitutive promoter) Plate 3: 19A – pB1A2 (V1013)
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#*BBa_R0062 (Top 10) (HSL and luxR inducible) Plate 1: 9G – pSB1A2 (V1004)
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#*BBa_J04500 (Top 10) (IPTG inducible prom + RBS) Plate 1: 16P – p5B1AK3 (V1009)
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#*BBa_E0040 (Top 10) (GFP mutant no promoter (3b)) Plate 1: 5H – p5B1A2 (V1001)
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#[[User:Mojs|Maia]] and [[User:Christinemerrick|Christine]] researched restriction enzymes for the reporter constructs and Mini-Tn5s streaked on 4/7/07 and digested some.  Had planned to make biobricks using Mini-Tn5s etc but discovered that a previous attempt to use transposable elements to make biobricks had been unsuccessful due to scarring at the restriction site. http://partsregistry.org/Part:BBa_J61206
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=== Friday 6 July 2007 ===
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{| align="center" border="1" cellpadding="5"
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|- align="center"
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|'''BioBrick Part''' || '''Function''' || '''Chassis''' || '''Plasmid''' || '''Kit Plate Location''' ''(Plate/Well)''
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|- align="center"
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|[http://partsregistry.org/Part:BBa_P1010 Bba_p1010] || Construction plasmid || ''E. coli''  DB3.1 || pSB1A10 || 4/7A
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|- align="center"
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| " || " || " || pSB1A10 || 4/11E
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|- align="center"
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| " || " || " || pSB3K3 || 4/11C
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|- align="center"
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|[http://partsregistry.org/Part:BBa_I52001 Bba_I52001] || High copy-number construction plasmid || ''E. coli''  TOP10 <br> <font color="red">should have been DB3.1</font> || pSB4A5 || 4/5I
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|- align="center"
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| " || " || " || pSB4K5 || 4/5D
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|- align="center"
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| " || " || " || pSB4K5 || 4/6D
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|- align="center"
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| " || " || " || pSB3K5 || 4/6B
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|- align="center"
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| " || " || " || pSB4A5 || 4/6E
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|- align="center"
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|[http://partsregistry.org/Part:BBa_J23119 Bba_J23119] || Strong constitutive promoter || ''E. coli''  TOP10 || pSB1A2 || 3/19A
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|- align="center"
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|[http://partsregistry.org/Part:BBa_R0062 Bba_R0062] || HSL & LuxR-inducible promoter || ''E. coli''  TOP10 || pSB1A2 || 1/9G
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|- align="center"
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|[http://partsregistry.org/Part:BBa_J04500 Bba_J04500] || IPTG-inducible promoter + RBS || ''E. coli''  TOP10 || pSB1AK3 || 1/16P
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|- align="center"
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|[http://partsregistry.org/Part:BBa_E0040 Bba_E0040] || Promoter-less GFP || ''E. coli''  TOP10 || pSB1A2 || 1/5H
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|}
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</li>
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<li>
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[[User:Mojs|Maia]] and [[User:Christinemerrick|Christine]] researched restriction enzymes for the reporter constructs and Mini-Tn5s streaked on 4/7/07 and digested some.  Had planned to make biobricks using Mini-Tn5s etc but discovered that [http://partsregistry.org/Part:BBa_J61206 a previous attempt to use transposable elements to make biobricks] had been unsuccessful due to scarring at the restriction site.
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</li>
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</ol>
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=== Friday 6th July 2007 ===
#[[User:MaijaP|Maija]] and [[User:Christinemerrick|Christine]] made 10x stocks for M9 (see [[Glasgow/Wetlab/Protocols#Protocol 3: Minimal Media and Trace Elements|Protocol 3.2 - M9]]).
#[[User:MaijaP|Maija]] and [[User:Christinemerrick|Christine]] made 10x stocks for M9 (see [[Glasgow/Wetlab/Protocols#Protocol 3: Minimal Media and Trace Elements|Protocol 3.2 - M9]]).
#Tutorial in [http://www.mbio.ncsu.edu/BioEdit/bioedit.html BioEdit] and [http://biotools.umassmed.edu/bioapps/primer3_www.cgi Primer3].
#Tutorial in [http://www.mbio.ncsu.edu/BioEdit/bioedit.html BioEdit] and [http://biotools.umassmed.edu/bioapps/primer3_www.cgi Primer3].
#Started designing primers for amplification and site directed mutagenesis of DmpR and DmpR #24 (see [[Glasgow/Wetlab/Protocols#Protocol 4: Primer Design|Protocol 4]]).
#Started designing primers for amplification and site directed mutagenesis of DmpR and DmpR #24 (see [[Glasgow/Wetlab/Protocols#Protocol 4: Primer Design|Protocol 4]]).
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<br>
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[[Glasgow/Wetlab/Week2|<font face=georgia color=#3366CC size=4>Next <br>  Week</font>]]
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----
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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 17:07, 22 October 2007

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Used 		Gels

Contents

Week 1

Tuesday 3rd July 2007

  1. Maija and Christine prepared LB broth and LB agar with Protocol 1. Made 4 batches of plates, 2 with kanamycin and 2 with carbenicillin.

Wednesday 4th July 2007

  1. All wetlab researched BioBricks.
  2. Reporter constructs and mini-Tn5 stocks looked out.
  3. Streaked the following:
    • P. putida PAW 340 pJAK14 (Carb. plate)
    • Tn5 lux AB (Carb. plate)
    • Mini-Tn5 lux AB (Carb. plate)
    • P. fluorescens NCIMB 9815 (Carb. plate)
    • P. putida KT2440 (LB)
    • JM109 pBluescript 5k+ (Carb. plate)
    • Mini-Tn5 Tc (Carb. Plate)
    • pQF52 (Carb. plate)
    • P. fluorescens 9815 (LB)
    • E. coli pJAK14 (Km plate)
    • Il DntR in pOF52 (Carb. plate)
    • pUCINR in Ω strain C (Carb. plate)
    • pGLTUR (Carb. plate)
    • Mini-Tn5 Kan (Carb. plate)
    • Mini-Tn5 Sm/Sp (Carb. plate)
    • Mini-Tn5 1cc2 (Carb. plate)
    • E. coli Sa1 (LB)
    • DmpR #24 (Carb. plate)
    • Mini-Tn5 lac 32 in E. coli 517 (Carb. plate)
    • Mini-Tn5 Tc (Carb. Plate)
    • Mini-Tn5 Cm (Carb. plate)
    • DmpR WT (Carb. plate)
    • E.coli sm 10 pESD15 Tn5 GFP (Carb. plate)
    • pUJ8 (Carb. Plate)

Thursday 5th July 2007

  1. BioBricks – Maija and Scott transformed using Protocol 2.
    BioBrick Part Function Chassis Plasmid Kit Plate Location (Plate/Well)
    [http://partsregistry.org/Part:BBa_P1010 Bba_p1010] Construction plasmid E. coli DB3.1 pSB1A10 4/7A
    " " " pSB1A10 4/11E
    " " " pSB3K3 4/11C
    [http://partsregistry.org/Part:BBa_I52001 Bba_I52001] High copy-number construction plasmid E. coli TOP10
    should have been DB3.1     		pSB4A5 4/5I
    " " " pSB4K5 4/5D
    " " " pSB4K5 4/6D
    " " " pSB3K5 4/6B
    " " " pSB4A5 4/6E
    [http://partsregistry.org/Part:BBa_J23119 Bba_J23119] Strong constitutive promoter E. coli TOP10 pSB1A2 3/19A
    [http://partsregistry.org/Part:BBa_R0062 Bba_R0062] HSL & LuxR-inducible promoter E. coli TOP10 pSB1A2 1/9G
    [http://partsregistry.org/Part:BBa_J04500 Bba_J04500] IPTG-inducible promoter + RBS E. coli TOP10 pSB1AK3 1/16P
    [http://partsregistry.org/Part:BBa_E0040 Bba_E0040] Promoter-less GFP E. coli TOP10 pSB1A2 1/5H
  2. Maia and Christine researched restriction enzymes for the reporter constructs and Mini-Tn5s streaked on 4/7/07 and digested some. Had planned to make biobricks using Mini-Tn5s etc but discovered that [http://partsregistry.org/Part:BBa_J61206 a previous attempt to use transposable elements to make biobricks] had been unsuccessful due to scarring at the restriction site.

Friday 6th July 2007

  1. Maija and Christine made 10x stocks for M9 (see Protocol 3.2 - M9).
  2. Tutorial in [http://www.mbio.ncsu.edu/BioEdit/bioedit.html BioEdit] and [http://biotools.umassmed.edu/bioapps/primer3_www.cgi Primer3].
  3. Started designing primers for amplification and site directed mutagenesis of DmpR and DmpR #24 (see Protocol 4).


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