Team:TorontoMaRSDiscovery/Notebook

From 2009.igem.org

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=April 27, 2009=
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[[image:To_igem_wiki_banner.jpg|965px]]
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#Received from Rosa (SPiT):
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{| style="color:white;background-color:#99CCFF;" height:100px cellpadding="2" cellspacing="0" border="0" width="100%" align="center" class="menu"
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#*TM0785
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!align="center"|[[Team:TorontoMaRSDiscovery|Home]]
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#**Plasmid containing encapsulin
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!align="center"|[[Team:TorontoMaRSDiscovery/Team|The Team]]
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#**Recommend transfect into bacteria and re-sequence
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!align="center"|[[Team:TorontoMaRSDiscovery/Project|The Project]]
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#**See email note regarding sequence error
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!align="center"|[[Team:TorontoMaRSDiscovery/Parts|BioBricks]]
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#*0.5 microliters TMG DNA 100 microgram/microliter
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!align="center"|[[Team:TorontoMaRSDiscovery/Modeling|Modelling]]
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#**Use 0.4 microliter for 50 microliter PCR reaction
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!align="center"|[[Team:TorontoMaRSDiscovery/Bioinformatics|Bioinformatics]]
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#**This is thumotoga maritime genomic DNA for purpose of re-cloning <Br />
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!align="center"|[[Team:TorontoMaRSDiscovery/Safety|Safety]]
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#Microcentrifuge tubes 1 and 2 placed in -20 freezer
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!align="center"|[[Team:TorontoMaRSDiscovery/Notebook|Notebook]]
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|}
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<br>
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=2009-05-15=
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==Monthly Notebook==
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<ul>
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<li>[https://2009.igem.org/Team:TorontoMaRSDiscovery/Notebook/April April]
 +
<li>[https://2009.igem.org/Team:TorontoMaRSDiscovery/Notebook/May May]
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<li>[https://2009.igem.org/Team:TorontoMaRSDiscovery/Notebook/June June]
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<li>[https://2009.igem.org/Team:TorontoMaRSDiscovery/Notebook/July July]
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<li>[https://2009.igem.org/Team:TorontoMaRSDiscovery/Notebook/Augst August]
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<li>[https://2009.igem.org/Team:TorontoMaRSDiscovery/Notebook/September September]
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<li>[https://2009.igem.org/Team:TorontoMaRSDiscovery/Notebook/October October]
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</ul>
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'''pH buffers received from VWR Mississauga'''
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==Summary of Results==
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*pH 4 buffer (red) 500 ml
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{| border="1" cellpadding="5" cellspacing="0"
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*pH 7 buffer (yellow) 500 ml
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|'''Part Type'''
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*pH 10 buffer (blue) 500 ml
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|'''Arbitrary Name'''
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|'''Registry Code'''
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''Above are used for pH/mV Meter calibration''  
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|'''Construct Used For'''
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|'''Status'''
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'''pH/mV meter calibrated according to manual – recorded in index'''  
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|'''Transformants Stocked?'''
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|'''Antibiotic Resistant, Backbone Plasmid Visualized?'''
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'''Ethanol solution (70%) made from 85% ethanol'''  
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|'''Part Visualized?'''
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|'''Sequenced?'''
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=2009-05-19=
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|----
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1. 2L of TE buffer made (10X TE)
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|rowspan="2"|
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a. Recipe for 2L from stock solution (10X TE)
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'''New Parts'''
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i. 10 ml 1M Tris-HCl pH 7.5-80.0 x 2 = 20 ml
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|Encapsulin (Enc)
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ii. 2 ml 0.5 M EDTA pH 8.0 x 2 = 4 ml
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|K192000
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iii. 988 ml ddH20 x 2 = 1976 ml
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|Encapsulin
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|Confirmed
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- To make Tris-HCl, Tris base is used and adjusted to desired pH using HCl
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|Yes
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- 1 M Tris base is required of which 20 ml is required – thereby 2.4228 g Tris base (due to scale limitations, 2.42g of Tris Base was used
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|Yes
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|Yes
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2. 500 ml of 1 M Tris Base made
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|Yes
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- mass of Tris used = 1M/1000 ml x 500 ml x 121.14 g/mol = 60.57 g
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|----
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- volume of water used = 500 ml
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|eCFPtgt
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|K192001
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3. 250 ml of 0.5 M EDTA solution was made
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|CFP target protein
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- mass of EDTA used = 36.53 g
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|Synthesized by Mr Gene
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- observations: EDTA did not dissolve in ddH2O on heat and being stirred
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|No
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|No
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10 MINUTE BRIEFING (INSERT)
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|No
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|Yes
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2009-05-21
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|----
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|rowspan="13"|
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- retrieved autoclaved ddH20, glycerol solution
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'''Registry Parts'''
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|1
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Gel Electrophoresis (test run)  
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|JJ23100
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|Control, Encapsulin
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- 1% gel: 0.5 g agar mixed with 50 ml of 1X TBE
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|Confirmed
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- 10X TBE diluted with 5 ml TBE + 45 ml ddH20 – did not use all 50 ml for 1 gel (gel will be too thick – make 2 instead)
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|Yes
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- Loading Dye: add glycerol solution to 25 mg bromophenol blue then add dH20 to 10 ml (to make 6X)  
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|Yes
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- Running gel: match wells to black side, run at 120 mA
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|Too small
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|No
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Visualize Gel in UV
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|----
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1. Turn power on
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|2
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2. Gel in machine face up
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|B0034
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3. Close door securely
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|Control, Encapsulin
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4. Turn white light on
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|Confirmed
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5. Adjust zoom, contrast, focus from black dial on top of machine
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|Yes
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6. Turn white light off (turns on UV)
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|Yes
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7. Press ‘live’ toggle – acq. Should be 0.4 sec.
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|Too small
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8. Print if desired or save on floppy disk
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|No
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9. Turn power off
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|----
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10. Dispose of gel in proper container
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|3
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11. Close door
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|C0040
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|Control
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2009-05-25
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|Confirmed
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|Yes
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- made overnight bacteria culture (200 ml LB: 20 microliters DB3.1)
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|Yes
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|Yes
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2009-05-26
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|No
 +
|----
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|4
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|C0012
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|Control
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|Confirmed
 +
|Yes
 +
|Yes
 +
|Yes
 +
|No
 +
|----
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|5
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|B0015
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|Control, Encapsulin
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|Confirmed
 +
|Yes
 +
|Yes
 +
|Yes
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|No
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|----
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|C (Amp + C)
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|pSB1AC3
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|Assembly
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|Confirmed
 +
|Yes
 +
|Yes
 +
|Yes
 +
|No
 +
|----
 +
|C (C only)
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|pSB1C3
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|Assembly
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|Confirmed
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|Yes
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|Yes
 +
|Yes
 +
|No
 +
|----
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|K (ccdb)
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|pSB1AK3
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|Assembly
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|Confirmed
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|Yes
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|Yes
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|Yes
 +
|No
 +
|----
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|K (RFP)
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|pSB1K3
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|Assembly
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|Confirmed
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|Yes
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|Yes
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|Yes
 +
|No
 +
|----
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|Tet (ccdb)
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|pSB1AT3
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|Assembly
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|Confirmed
 +
|Yes
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|Yes
 +
|Yes
 +
|No
 +
|----
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|Tet (RFP)
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|pSB1T3
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|Assembly
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|Confirmed
 +
|Yes
 +
|Yes
 +
|Yes
 +
|No
 +
|----
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|7
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|J13002
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|Encapsulin
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|Transfected
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|Yes
 +
|No
 +
|No
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|No
 +
|----
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|Amp
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|pSB1A3
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|Assembly
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|Transfected
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|Yes
 +
|No
 +
|No
 +
|No
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|-
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|rowspan="7"|
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'''Assembled Parts'''
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|----
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|Enc in C (Amp+C)
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|
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|Submission, Encapsulin
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|Confirmed
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|Yes
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|Yes
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|Yes
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|Yes
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|----
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|1+2 in C (Amp+C)
 +
|
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|Control, Encapsulin
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|Confirmed
 +
|Yes
 +
|Yes
 +
|Too small
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|No
 +
|----
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|3+2 in C (Amp+C)
 +
|
 +
|Control
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|Confirmed
 +
|Yes
 +
|Yes
 +
|Yes
 +
|No
 +
|----
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|4+5 in Tet (ccdb)
 +
|
 +
|Control
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|Confirmed
 +
|Yes
 +
|Yes
 +
|Yes
 +
|No
 +
|----
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|1+2+3+2 in K (RFP)
 +
|
 +
|Control
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|Transfected
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|Yes
 +
|No
 +
|No
 +
|No
 +
|----
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|1+2+Enc in K
 +
|
 +
|Encapsulin
 +
|Confirmed
 +
|Yes
 +
|Yes
 +
|Yes
 +
|In Progress
 +
|----
 +
|}

Latest revision as of 03:05, 22 October 2009

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Home The Team The Project BioBricks Modelling Bioinformatics Safety Notebook


Monthly Notebook

Summary of Results

Part Type Arbitrary Name Registry Code Construct Used For Status Transformants Stocked? Antibiotic Resistant, Backbone Plasmid Visualized? Part Visualized? Sequenced?

New Parts

Encapsulin (Enc) K192000 Encapsulin Confirmed Yes Yes Yes Yes
eCFPtgt K192001 CFP target protein Synthesized by Mr Gene No No No Yes

Registry Parts

1 JJ23100 Control, Encapsulin Confirmed Yes Yes Too small No
2 B0034 Control, Encapsulin Confirmed Yes Yes Too small No
3 C0040 Control Confirmed Yes Yes Yes No
4 C0012 Control Confirmed Yes Yes Yes No
5 B0015 Control, Encapsulin Confirmed Yes Yes Yes No
C (Amp + C) pSB1AC3 Assembly Confirmed Yes Yes Yes No
C (C only) pSB1C3 Assembly Confirmed Yes Yes Yes No
K (ccdb) pSB1AK3 Assembly Confirmed Yes Yes Yes No
K (RFP) pSB1K3 Assembly Confirmed Yes Yes Yes No
Tet (ccdb) pSB1AT3 Assembly Confirmed Yes Yes Yes No
Tet (RFP) pSB1T3 Assembly Confirmed Yes Yes Yes No
7 J13002 Encapsulin Transfected Yes No No No
Amp pSB1A3 Assembly Transfected Yes No No No

Assembled Parts

Enc in C (Amp+C) Submission, Encapsulin Confirmed Yes Yes Yes Yes
1+2 in C (Amp+C) Control, Encapsulin Confirmed Yes Yes Too small No
3+2 in C (Amp+C) Control Confirmed Yes Yes Yes No
4+5 in Tet (ccdb) Control Confirmed Yes Yes Yes No
1+2+3+2 in K (RFP) Control Transfected Yes No No No
1+2+Enc in K Encapsulin Confirmed Yes Yes Yes In Progress
Retrieved from "http://2009.igem.org/Team:TorontoMaRSDiscovery/Notebook"