Melbourne/Lab Notebook

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[[Melbourne|<Back to team home page>]]  
[[Melbourne|<Back to team home page>]]  
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*Gas Vesicle lab notebook:
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** [[Melbourne/Lab GV Notebook|Gas Vesicles Notebook]]
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==Week 1==
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*Blue photoreceptor notebook:
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*25 June 2007: Prepared LB agar plates Amp & Kana.
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** [[Melbourne/Lab BL Notebook|Blue Photoreceptor Notebook]]
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*25 June 2007: [[Melbourne/IGEM 2007 kit|Resuspended the following from Registry plates]] & [[Melbourne/Transformation Protocol shorter|Transformed (shorter protocol)]] into competent DH5alpha cells.
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*#[[Melbourne/BBa_I15008|'''P2 21A''']] - (BBa_I15008, ho1, Kan) -> No colonies on plates
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*#[[Melbourne/BBa_I15009|'''P2 21C''']] - (BBa_I15009, PcyA, Kan) ->No colonies on plates
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*#[[Melbourne/BBa_R0084|'''P1 11H''']] - (BBa_R0084, OmpR positive promoter, Amp)->Small number of colonies.
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*Photoreceptor & control system lab notebooks:
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** [[Melbourne/Lab Notebook Weeks 1-4|Weeks 1-4]]
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*26 June 2007: [[Melbourne/IGEM 2007 kit|Resuspended the following from Registry plates]] & [[Melb:Transformation Protocol|Transformed]] into Joe's competent DH5alpha cells.
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** [[Melbourne/Lab Notebook Weeks 5-8|Weeks 5-8]]
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*#[[Melbourne/BBa_B0034|'''P1 3O''']] (BBa_B0034, RBS, Amp)
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** [[Melbourne/Lab Notebook Weeks 9-12|Weeks 9-12]]
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*#[[Melbourne/BBa_C0051|'''P1 5G''']] (BBa_C0051, c1 protein, Amp)
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** [[Melbourne/Lab Notebook Weeks 13-16|Weeks 13-16]]
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*#[[Melbourne/BBa_B0010|'''P2 3P''']] (BBa_B0010, Terminator, Amp)
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** [[Melbourne/Lab Notebook Weeks 17-18|Weeks 17-18]]
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*26 June 2007: Streaked the following cells:
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*#[[Melbourne/pJS010|'''pJS010''']] (from solid agar, Amp)
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*#[[Melbourne/Fusion|'''Fusion protein''']] (from glycerol stock, Amp?)
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*#[[Melbourne/BBa_I15010|'''BBa_I15010''']] (from solid agar, Kan)
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*27 June 2007: [[Melb:Transformation Protocol|Transformed]] into Joe's competent DH5alpha cells.
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*#[[Melbourne/BBa_I15008|'''P2 21A''']] (Kan)
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*#[[Melbourne/BBa_I15009|'''P2 21C''']] (Kan)
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====Liquid culture====
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#[[Melb:Growing up cells|Prepared]] 30mL of Amp LB (100ug/mL). Prepared 5mL Kan LB (50mg/mL).
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#Aliquoted 5mL Amp LB into 6 50mL falcon tubes
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#To the Amp LB aliquots single transformed colonies from plates of the following were introduced:
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#*[[Melbourne/pJS010|'''pJS010''']]
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#*[[Melbourne/Fusion|'''Fusion''']]
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#*[[Melbourne/BBa_B0034|'''P1 3O''']]
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#*[[Melbourne/BBa_C0051|'''P1 5G''']]
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#*[[Melbourne/BBa_B0010|'''P2 3P''']]
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#*[[Melbourne/BBa_R0084|'''P1 11H''']]
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#To the Kan LB a single colony from the transformation plate of [[Melbourne/BBa_I15010|'''BBa_I15010''']] was introduced.
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#Cells incubated at 37degrees with shaking overnight.
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===28 June 2007===
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====Miniprep====
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#[[Melb:Miniprep protocol|Minipreped]] the cultures grown overnight after putting aside 1mL(sterile) for glycerol stocks:
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#*[[Melbourne/pJS010|'''pJS010''']]
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#*[[Melbourne/Fusion|'''Fusion''']]
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#*[[Melbourne/BBa_B0034|'''P1 3O''']]
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#*[[Melbourne/BBa_C0051|'''P1 5G''']]
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#*[[Melbourne/BBa_B0010|'''P2 3P''']]
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#*[[Melbourne/BBa_R0084|'''P1 11H''']]
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#*[[Melbourne/BBa_I15010|'''I15010''']]
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#Stored in -20 freezer
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====Digest====
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====Liquid culture====
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#[[Melb:Growing up cells|Cultured]] the following cells from transformed plates:
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#*[[Melbourne/BBa_R0084|'''P1 11H''']] (BBa_R0084)
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#*[[Melbourne/BBa_I15010|'''BBa_I15010''']]
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#*[[Melbourne/BBa_B0034|'''P1 3O''']] (BBa_B0034)
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#*[[Melbourne/BBa_C0051|'''P1 5G''']] (BBa_C0051)
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#*[[Melbourne/BBa_I15008|'''P1 21A''']] (BBa_I15008)
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#*[[Melbourne/BBa_I15009|'''P1 21C''']] (BBa_I15009)
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===29 June 2007===
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====Miniprep====
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#[[Melb:Miniprep protocol|Minipreped]] the cultures grown overnight after putting aside 1mL(sterile) for glycerol stocks:
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#*[[Melbourne/BBa_R0084|'''P1 11H''']] (BBa_R0084)
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#*[[Melbourne/BBa_I15010|'''BBa_I15010''']]
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#*[[Melbourne/BBa_B0034|'''P1 3O''']] (BBa_B0034)
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#*[[Melbourne/BBa_C0051|'''P1 5G''']] (BBa_C0051)
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#*[[Melbourne/BBa_I15008|'''P1 21A''']] (BBa_I15008)
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#*[[Melbourne/BBa_I15009|'''P1 21C''']] (BBa_I15009)
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#Stored in -20 freezer
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===30 June 2007===
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==Week 2==
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*2 July 2007: [[Melbourne/IGEM 2007 kit|Resuspended the following from Registry plates]] & [[Melb:Transformation Protocol|Transformed]] into Joe's competent DH5alpha cells.
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*#Q04510
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*#E0241
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*#E0040
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*#B0014
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*#J61035
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*3 July 2007: [[Melb:Transformation Protocol|Re Transformed]] into Joe's competent DH5alpha cells. Each Plate had additional 40uL of 50mg/ml Ampicillan spread on surface.
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*#[[Melbourne/BBa_J61035|'''P4 8J''']]  -> Three colonies -> grew in liquid culture 4 july
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*#[[Melbourne/BBa_E0040|'''P1 5H''']] -> Multiple colonies at edge -> did not grow in overnight liquid culture 4 july 'poor amp spreading'
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*#[[Melbourne/BBa_E0241|'''P2 15L''']] -> Three colonies -> grew in liquid culture 4 july
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*4 July 2007:
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====Ampicillin Plates====
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*Made up 25 Ampicillin plates using 400mL LB agar prepared on the 25th of June. 
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**Used 100mg/ml Ampicillin stock from Gooley Lab - In bag in left freezer near our lab bench.
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====Tranformation====
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[[Melbourne/Transformation Protocol|Transformed]] the following and grew on new ampicillin plates
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*[[Melbourne/BBa_E0040|'''P1 5H''']]
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*[[Melbourne/BBa_J61035|'''P4 8J''']]
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*[[Melbourne/BBa_E0241|'''P2 15L''']]
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**Also Plated an untransformed control (subjected to transformation protocol in absense of DNA) and a blank plate control (no cells)
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====Liquid Culture====
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*[[Melbourne/Growing up cells|Cultured]] 2 colonies from each of the following transformed plates and labelled as follows
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**[[Melbourne/BBa_B0010|'''P2 3P 1''']]
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**[[Melbourne/BBa_B0010|'''P2 3P 2''']]
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**[[Melbourne/BBa_I15010|'''I15010 1''']]
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**[[Melbourne/BBa_I15010|'''I15010 2''']]
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**[[Melbourne/pJS010|'''pJS010 1''']]
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**[[Melbourne/pJS010|'''pJS010 2''']]
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**[[Melbourne/Fusion|'''Fusion 1''']]
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**[[Melbourne/Fusion|'''Fusion 2''']]
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**[[Melbourne/BBa_J61035|'''P4 8J 1''']]
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**[[Melbourne/BBa_J61035|'''P4 8J 2''']]
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**[[Melbourne/BBa_E0241|'''P2 15L 1''']]
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**[[Melbourne/BBa_E0241|'''P2 15L 2''']]
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**[[Melbourne/BBa_Q04510|'''P2 13K 1''']]
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**[[Melbourne/BBa_Q04510|'''P2 13K 2''']]
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**[[Melbourne/BBa_B0014|'''P1 1G 1''']]
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**[[Melbourne/BBa_B0014|'''P1 1G 2''']]
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**[[Melbourne/BBa_R0084|'''P1 11H 1''']]
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**[[Melbourne/BBa_R0084|'''P1 11H 2''']]
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**[[Melbourne/BBa_E0040|'''P1 5H 1''']]
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**[[Melbourne/BBa_E0040|'''P1 5H 2''']]
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**suspected contaminating colony from [[Melbourne/BBa_J61035|'''P4 8J''']]
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===5 July 2007===
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====Miniprep====
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*1mL culture from the 4th of Julyput aside for glycerol stocks under sterile conditions.  Labelled with todays date 5/7
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*miniprepped the following overnight cultures set up on the 4th of July.  Final elution was performed with TE buffer prepared on 25/06/07 rather than nuclease free water. Labelled with todays date 5/7.
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**[[Melbourne/BBa_B0010|'''P2 3P 1''']]
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**[[Melbourne/BBa_B0010|'''P2 3P 2''']]
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**[[Melbourne/BBa_I15010|'''I15010 1''']]
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**[[Melbourne/BBa_I15010|'''I15010 2''']]
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**[[Melbourne/pJS010|'''pJS010 1''']]
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**[[Melbourne/pJS010|'''pJS010 2''']]
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**[[Melbourne/Fusion|'''Fusion 1''']]
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**[[Melbourne/Fusion|'''Fusion 2''']]
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**[[Melbourne/BBa_J61035|'''P4 8J 1''']]
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**[[Melbourne/BBa_J61035|'''P4 8J 2''']]
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**[[Melbourne/BBa_E0241|'''P2 15L 1''']]
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**[[Melbourne/BBa_E0241|'''P2 15L 2''']]
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**[[Melbourne/BBa_Q04510|'''P2 13K 1''']](eluted in 130uL due to accidental double application of 50uL elution)
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**[[Melbourne/BBa_Q04510|'''P2 13K 2''']]
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**[[Melbourne/BBa_B0014|'''P1 1G 1''']]
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**[[Melbourne/BBa_B0014|'''P1 1G 2''']]
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*the following liquid cultures were not miniprepped due to failure (no growth)
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**[[Melbourne/BBa_R0084|'''P1 11H 1''']]
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**[[Melbourne/BBa_R0084|'''P1 11H 2''']]
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**[[Melbourne/BBa_E0040|'''P1 5H 1''']]
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**[[Melbourne/BBa_E0040|'''P1 5H 2''']]
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**suspected contaminating colony from [[Melbourne/BBa_J61035|'''P4 8J''']]
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====Digest====
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Performed the following digests on DNA from the above miniprep
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=====EcoRI/PstI with buffer 3=====
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*[[Melbourne/BBa_I15010|'''I15010 1''']]
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*[[Melbourne/BBa_I15010|'''I15010 2''']]
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*[[Melbourne/BBa_J61035|'''P4 8J 1''']]
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*[[Melbourne/BBa_J61035|'''P4 8J 2''']]
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*[[Melbourne/BBa_E0241|'''P2 15L 1''']]
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*[[Melbourne/BBa_E0241|'''P2 15L 2''']]
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*[[Melbourne/BBa_Q04510|'''P2 13K 1''']]
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*[[Melbourne/BBa_Q04510|'''P2 13K 2''']]
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=====EcoRI/HaeII in buffer 2=====
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*[[Melbourne/BBa_B0010|'''P2 3P 1''']]
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*[[Melbourne/BBa_B0010|'''P2 3P 2''']]
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*[[Melbourne/BBa_B0014|'''P1 1G 1''']]
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*[[Melbourne/BBa_B0014|'''P1 1G 2''']]
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=====XbaI/SpeI in buffer 2=====
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*[[Melbourne/pJS010|'''pJS010 1''']]
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*[[Melbourne/pJS010|'''pJS010 2''']]
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Incubated for 2hours at 37 degrees, before addition of loading dye and storage at -20
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====Transformation====
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*[[Melbourne/Transformation Protocol|Transformed]] P1 11H from resuspended DNA.
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====Liquid Culture====
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*[[Melbourne/BBa_E0040|'''P1 5H 1''']]
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*[[Melbourne/BBa_E0040|'''P1 5H 2''']]
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*[[Melbourne/BBa_J61035|'''P4 8J 1''']]
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*[[Melbourne/BBa_J61035|'''P4 8J 2''']]
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*[[Melbourne/BBa_E0241|'''P2 15L 1''']]
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*[[Melbourne/BBa_E0241|'''P2 15L 2''']]
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===6 July 2007===
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====Digest Gel====
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* Prepared 20 lane 100mL [[Melbourne/Preparing an agarose gel|agarose gel]] with 0.5xTBE buffer.
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*Loaded 20uL of digest samples in the following lane order
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*#1kb+ ladder
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*Ran for 1.5hours
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====Miniprep====
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*Put aside 1mL of liquid cultures set up on the 5th for glycerol stocks and labelled with todays date 6/7
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*Miniprepped the remains of the cultures and labelled with todays date 6/7.  Samples were eluted with TE buffer.
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*[[Melbourne/BBa_E0040|'''P1 5H 1''']]
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*[[Melbourne/BBa_E0040|'''P1 5H 2''']]
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*[[Melbourne/BBa_J61035|'''P4 8J 1''']]
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*[[Melbourne/BBa_J61035|'''P4 8J 2''']]
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*[[Melbourne/BBa_E0241|'''P2 15L 1''']]
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*[[Melbourne/BBa_E0241|'''P2 15L 2''']]
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====Digest====
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Digested
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==Week 3==
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===9 July 2007===
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===10 July 2007===
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===11 July 2007===
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===12 July 2007===
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===13 July 2007===
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==Week 4==
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===16 July 2007===
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===17 July 2007===
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===18 July 2007===
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===19 July 2007===
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===20 July 2007===
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==Week 5==
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===23 July 2007===
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===24 July 2007===
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===25 July 2007===
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===26 July 2007===
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===27 July 2007===
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==Week 6==
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===30 July 2007===
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===31 July 2007===
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===1  Aug 2007===
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===2  Aug 2007===
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===3  Aug 2007===
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Latest revision as of 14:25, 26 October 2007

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