Melbourne/Lab Notebook

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[[Melbourne|<Back to team home page>]]  
[[Melbourne|<Back to team home page>]]  
 +
*Gas Vesicle lab notebook:
 +
** [[Melbourne/Lab GV Notebook|Gas Vesicles Notebook]]
-
==Week 1==
+
*Blue photoreceptor notebook:
-
*25 June 2007: Prepared LB agar plates Amp & Kana.
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** [[Melbourne/Lab BL Notebook|Blue Photoreceptor Notebook]]
-
*25 June 2007: [[Melbourne/IGEM 2007 kit|Resuspended the following from Registry plates]] & [[Melbourne/Transformation Protocol shorter|Transformed (shorter protocol)]] into competent DH5alpha cells.
+
-
*#[[Melbourne/BBa_I15008|'''P2 21A''']] - (BBa_I15008, ho1, Kan) -> No colonies on plates
+
-
*#[[Melbourne/BBa_I15009|'''P2 21C''']] - (BBa_I15009, PcyA, Kan) ->No colonies on plates
+
-
*#[[Melbourne/BBa_R0084|'''P1 11H''']] - (BBa_R0084, OmpR positive promoter, Amp)->Small number of colonies.
+
-
 
+
*Photoreceptor & control system lab notebooks:
-
 
+
** [[Melbourne/Lab Notebook Weeks 1-4|Weeks 1-4]]
-
*26 June 2007: [[Melbourne/IGEM 2007 kit|Resuspended the following from Registry plates]] & [[Melb:Transformation Protocol|Transformed]] into Joe's competent DH5alpha cells.
+
** [[Melbourne/Lab Notebook Weeks 5-8|Weeks 5-8]]
-
*#[[Melbourne/BBa_B0034|'''P1 3O''']] (BBa_B0034, RBS, Amp)
+
** [[Melbourne/Lab Notebook Weeks 9-12|Weeks 9-12]]
-
*#[[Melbourne/BBa_C0051|'''P1 5G''']] (BBa_C0051, c1 protein, Amp)
+
** [[Melbourne/Lab Notebook Weeks 13-16|Weeks 13-16]]
-
*#[[Melbourne/BBa_B0010|'''P2 3P''']] (BBa_B0010, Terminator, Amp)
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** [[Melbourne/Lab Notebook Weeks 17-18|Weeks 17-18]]
-
 
+
-
*26 June 2007: Streaked the following cells:
+
-
*#[[Melbourne/pJS010|'''pJS010''']] (from solid agar, Amp)
+
-
*#[[Melbourne/Fusion|'''Fusion protein''']] (from glycerol stock, Amp?)
+
-
*#[[Melbourne/BBa_I15010|'''BBa_I15010''']] (from solid agar, Kan)
+
-
 
+
-
*27 June 2007: [[Melb:Transformation Protocol|Transformed]] into Joe's competent DH5alpha cells.
+
-
*#[[Melbourne/BBa_I15008|'''P2 21A''']] (Kan)
+
-
*#[[Melbourne/BBa_I15009|'''P2 21C''']] (Kan)
+
-
 
+
-
 
+
-
====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''']]
+
-
#*[[Melbourne/BBa_B0034|'''P1 3O''']]
+
-
#*[[Melbourne/BBa_C0051|'''P1 5G''']]
+
-
#*[[Melbourne/BBa_B0010|'''P2 3P''']]
+
-
#*[[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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-
 
+
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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:
+
-
#*[[Melbourne/pJS010|'''pJS010''']]
+
-
#*[[Melbourne/Fusion|'''Fusion''']]
+
-
#*[[Melbourne/BBa_B0034|'''P1 3O''']]
+
-
#*[[Melbourne/BBa_C0051|'''P1 5G''']]
+
-
#*[[Melbourne/BBa_B0010|'''P2 3P''']]
+
-
#*[[Melbourne/BBa_R0084|'''P1 11H''']]
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-
#*[[Melbourne/BBa_I15010|'''I15010''']]
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-
#Stored in -20 freezer
+
-
 
+
-
====Digest====
+
-
 
+
-
 
+
-
====Liquid culture====
+
-
#[[Melb:Growing up cells|Cultured]] the following cells from transformed plates:
+
-
#*[[Melbourne/BBa_R0084|'''P1 11H''']] (BBa_R0084)
+
-
#*[[Melbourne/BBa_I15010|'''BBa_I15010''']]
+
-
#*[[Melbourne/BBa_B0034|'''P1 3O''']] (BBa_B0034)
+
-
#*[[Melbourne/BBa_C0051|'''P1 5G''']] (BBa_C0051)
+
-
#*[[Melbourne/BBa_I15008|'''P1 21A''']] (BBa_I15008)
+
-
#*[[Melbourne/BBa_I15009|'''P1 21C''']] (BBa_I15009)
+
-
 
+
-
===29 June 2007===
+
-
====Miniprep====
+
-
#[[Melb:Miniprep protocol|Minipreped]] the cultures grown overnight after putting aside 1mL(sterile) for glycerol stocks:
+
-
#*[[Melbourne/BBa_R0084|'''P1 11H''']] (BBa_R0084)
+
-
#*[[Melbourne/BBa_I15010|'''BBa_I15010''']]
+
-
#*[[Melbourne/BBa_B0034|'''P1 3O''']] (BBa_B0034)
+
-
#*[[Melbourne/BBa_C0051|'''P1 5G''']] (BBa_C0051)
+
-
#*[[Melbourne/BBa_I15008|'''P1 21A''']] (BBa_I15008)
+
-
#*[[Melbourne/BBa_I15009|'''P1 21C''']] (BBa_I15009)
+
-
 
+
-
#Stored in -20 freezer
+
-
 
+
-
===30 June 2007===
+
-
==Week 2==
+
-
*2 July 2007: [[Melbourne/IGEM 2007 kit|Resuspended the following from Registry plates]] & [[Melb:Transformation Protocol|Transformed]] into Joe's competent DH5alpha cells.
+
-
*#Q04510
+
-
*#E0241
+
-
*#E0040
+
-
*#B0014
+
-
*#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.
+
-
*#[[Melbourne/BBa_J61035|'''P4 8J''']]  -> Three colonies -> grew in liquid culture 4 july
+
-
*#[[Melbourne/BBa_E0040|'''P1 5H''']] -> Multiple colonies at edge -> did not grow in overnight liquid culture 4 july 'poor amp spreading'
+
-
*#[[Melbourne/BBa_E0241|'''P2 15L''']] -> Three colonies -> grew in liquid culture 4 july
+
-
 
+
-
*4 July 2007:
+
-
 
+
-
====Ampicillin Plates====
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-
*Made up 25 Ampicillin plates using 400mL LB agar prepared on the 25th of June. 
+
-
**Used 100mg/ml Ampicillin stock from Gooley Lab - In bag in left freezer near our lab bench.
+
-
 
+
-
====Tranformation====
+
-
[[Melbourne/Transformation Protocol|Transformed]] the following and grew on new ampicillin plates
+
-
*[[Melbourne/BBa_E0040|'''P1 5H''']]
+
-
*[[Melbourne/BBa_J61035|'''P4 8J''']]
+
-
*[[Melbourne/BBa_E0241|'''P2 15L''']]
+
-
**Also Plated an untransformed control (subjected to transformation protocol in absense of DNA) and a blank plate control (no cells)
+
-
 
+
-
====Liquid Culture====
+
-
*[[Melbourne/Growing up cells|Cultured]] 2 colonies from each of the following transformed plates and labelled as follows
+
-
**[[Melbourne/BBa_B0010|'''P2 3P 1''']]
+
-
**[[Melbourne/BBa_B0010|'''P2 3P 2''']]
+
-
**[[Melbourne/BBa_I15010|'''I15010 1''']]
+
-
**[[Melbourne/BBa_I15010|'''I15010 2''']]
+
-
**[[Melbourne/pJS010|'''pJS010 1''']]
+
-
**[[Melbourne/pJS010|'''pJS010 2''']]
+
-
**[[Melbourne/Fusion|'''Fusion 1''']]
+
-
**[[Melbourne/Fusion|'''Fusion 2''']]
+
-
**[[Melbourne/BBa_J61035|'''P4 8J 1''']]
+
-
**[[Melbourne/BBa_J61035|'''P4 8J 2''']]
+
-
**[[Melbourne/BBa_E0241|'''P2 15L 1''']]
+
-
**[[Melbourne/BBa_E0241|'''P2 15L 2''']]
+
-
**[[Melbourne/BBa_Q04510|'''P2 13K 1''']]
+
-
**[[Melbourne/BBa_Q04510|'''P2 13K 2''']]
+
-
**[[Melbourne/BBa_B0014|'''P1 1G 1''']]
+
-
**[[Melbourne/BBa_B0014|'''P1 1G 2''']]
+
-
**[[Melbourne/BBa_R0084|'''P1 11H 1''']]
+
-
**[[Melbourne/BBa_R0084|'''P1 11H 2''']]
+
-
**[[Melbourne/BBa_E0040|'''P1 5H 1''']]
+
-
**[[Melbourne/BBa_E0040|'''P1 5H 2''']]
+
-
**suspected contaminating colony from [[Melbourne/BBa_J61035|'''P4 8J''']]
+
-
 
+
-
===5 July 2007===
+
-
 
+
-
====Miniprep====
+
-
*1mL culture from the 4th of Julyput aside for glycerol stocks under sterile conditions.  Labelled with todays date 5/7
+
-
*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.
+
-
**[[Melbourne/BBa_B0010|'''P2 3P 1''']]
+
-
**[[Melbourne/BBa_B0010|'''P2 3P 2''']]
+
-
**[[Melbourne/BBa_I15010|'''I15010 1''']]
+
-
**[[Melbourne/BBa_I15010|'''I15010 2''']]
+
-
**[[Melbourne/pJS010|'''pJS010 1''']]
+
-
**[[Melbourne/pJS010|'''pJS010 2''']]
+
-
**[[Melbourne/Fusion|'''Fusion 1''']]
+
-
**[[Melbourne/Fusion|'''Fusion 2''']]
+
-
**[[Melbourne/BBa_J61035|'''P4 8J 1''']]
+
-
**[[Melbourne/BBa_J61035|'''P4 8J 2''']]
+
-
**[[Melbourne/BBa_E0241|'''P2 15L 1''']]
+
-
**[[Melbourne/BBa_E0241|'''P2 15L 2''']]
+
-
**[[Melbourne/BBa_Q04510|'''P2 13K 1''']](eluted in 130uL due to accidental double application of 50uL elution)
+
-
**[[Melbourne/BBa_Q04510|'''P2 13K 2''']]
+
-
**[[Melbourne/BBa_B0014|'''P1 1G 1''']]
+
-
**[[Melbourne/BBa_B0014|'''P1 1G 2''']]
+
-
*the following liquid cultures were not miniprepped due to failure (no growth)
+
-
**[[Melbourne/BBa_R0084|'''P1 11H 1''']]
+
-
**[[Melbourne/BBa_R0084|'''P1 11H 2''']]
+
-
**[[Melbourne/BBa_E0040|'''P1 5H 1''']]
+
-
**[[Melbourne/BBa_E0040|'''P1 5H 2''']]
+
-
**suspected contaminating colony from [[Melbourne/BBa_J61035|'''P4 8J''']]
+
-
 
+
-
====Digest====
+
-
Performed the following [[Melbourne/Diagnostic Digest|digests]] on DNA from the above miniprep
+
-
=====EcoRI/PstI with buffer 3=====
+
-
*[[Melbourne/BBa_I15010|'''I15010 1''']]
+
-
*[[Melbourne/BBa_I15010|'''I15010 2''']]
+
-
*[[Melbourne/BBa_J61035|'''P4 8J 1''']]
+
-
*[[Melbourne/BBa_J61035|'''P4 8J 2''']]
+
-
*[[Melbourne/BBa_E0241|'''P2 15L 1''']]
+
-
*[[Melbourne/BBa_E0241|'''P2 15L 2''']]
+
-
*[[Melbourne/BBa_Q04510|'''P2 13K 1''']]
+
-
*[[Melbourne/BBa_Q04510|'''P2 13K 2''']]
+
-
=====EcoRI/HaeII in buffer 2=====
+
-
*[[Melbourne/BBa_B0010|'''P2 3P 1''']]
+
-
*[[Melbourne/BBa_B0010|'''P2 3P 2''']]
+
-
*[[Melbourne/BBa_B0014|'''P1 1G 1''']]
+
-
*[[Melbourne/BBa_B0014|'''P1 1G 2''']]
+
-
=====XbaI/SpeI in buffer 2=====
+
-
*[[Melbourne/pJS010|'''pJS010 1''']]
+
-
*[[Melbourne/pJS010|'''pJS010 2''']]
+
-
 
+
-
Incubated for 2hours at 37 degrees, before addition of loading dye and storage at -20
+
-
 
+
-
====Transformation====
+
-
*[[Melbourne/Transformation Protocol|Transformed]] P1 11H from resuspended DNA.
+
-
 
+
-
====Liquid Culture====
+
-
*[[Melbourne/BBa_E0040|'''P1 5H 1''']]
+
-
*[[Melbourne/BBa_E0040|'''P1 5H 2''']]
+
-
*[[Melbourne/BBa_J61035|'''P4 8J 1''']]
+
-
*[[Melbourne/BBa_J61035|'''P4 8J 2''']]
+
-
*[[Melbourne/BBa_E0241|'''P2 15L 1''']]
+
-
*[[Melbourne/BBa_E0241|'''P2 15L 2''']]
+
-
 
+
-
===6 July 2007===
+
-
 
+
-
====Digest Gel====
+
-
* Prepared 20 lane 100mL [[Melbourne/Preparing an agarose gel|agarose gel]] with 0.5xTBE buffer.
+
-
*[[Melbourne/Loading a DNA gel|Loaded]] 20uL of digest samples in the following lane order
+
-
*#1kb+ ladder
+
-
*#[[Melbourne/BBa_I15010|'''I15010 1''']]
+
-
*#[[Melbourne/BBa_I15010|'''I15010 2''']]
+
-
*#[[Melbourne/BBa_Q04510|'''P2 13K 1''']]
+
-
*#[[Melbourne/BBa_Q04510|'''P2 13K 2''']]
+
-
*#[[Melbourne/BBa_E0241|'''P2 15L 1''']]
+
-
*#[[Melbourne/BBa_E0241|'''P2 15L 2''']]
+
-
*#[[Melbourne/BBa_J61035|'''P4 8J 1''']]
+
-
*#[[Melbourne/BBa_J61035|'''P4 8J 2''']]
+
-
*#[[Melbourne/BBa_B0014|'''P1 1G 1''']]
+
-
*#[[Melbourne/BBa_B0014|'''P1 1G 2''']]
+
-
*#[[Melbourne/BBa_B0010|'''P2 3P 1''']]
+
-
*#[[Melbourne/BBa_B0010|'''P2 3P 2''']]
+
-
*#[[Melbourne/pJS010|'''pJS010 1''']]
+
-
*#[[Melbourne/pJS010|'''pJS010 2''']]
+
-
*Ran for 1.5hours at 95V
+
-
 
+
-
====Miniprep====
+
-
*Put aside 1mL of liquid cultures set up on the 5th for glycerol stocks and labelled with todays date 6/7
+
-
*Miniprepped the remains of the cultures and labelled with todays date 6/7.  Samples were eluted with TE buffer.
+
-
*[[Melbourne/BBa_E0040|'''P1 5H 1''']]
+
-
*[[Melbourne/BBa_E0040|'''P1 5H 2''']]
+
-
*[[Melbourne/BBa_J61035|'''P4 8J 1''']]
+
-
*[[Melbourne/BBa_J61035|'''P4 8J 2''']]
+
-
*[[Melbourne/BBa_E0241|'''P2 15L 1''']]
+
-
*[[Melbourne/BBa_E0241|'''P2 15L 2''']]
+
-
 
+
-
 
+
-
====Digest====
+
-
*[[Melbourne/Diagnostic Digest|Digested]] 5uL of each of the above miniprep DNA with EcoRI and PstI in buffer 3.
+
-
*Incubated for 2hours 25min at 37degrees
+
-
*Added 5uL 6x loading dye and stored at -20
+
-
 
+
-
====Glycerol Stocks====
+
-
The following [[Melbourne/Glycerol Stocks|glycerol stocks]] were made:
+
-
*Put aside from cultures 5/7 (labelled with this date)
+
-
*#[[Melbourne/BBa_E0241|'''P2 15L 1''']]
+
-
*#[[Melbourne/BBa_E0241|'''P2 15L 2''']]
+
-
*#[[Melbourne/BBa_J61035|'''P4 8J 1''']]
+
-
*#[[Melbourne/BBa_J61035|'''P4 8J 2''']]
+
-
*#[[Melbourne/BBa_B0014|'''P1 1G 1''']]
+
-
*#[[Melbourne/BBa_B0014|'''P1 1G 2''']]
+
-
*Put aside from cultures 6/7 (labelled with this date)
+
-
*#[[Melbourne/BBa_E0241|'''P2 15L 1''']]
+
-
*#[[Melbourne/BBa_E0241|'''P2 15L 2''']]
+
-
*#[[Melbourne/BBa_J61035|'''P4 8J 1''']]
+
-
*#[[Melbourne/BBa_J61035|'''P4 8J 2''']]
+
-
*[[Melbourne/BBa_E0040|'''P1 5H 1''']]
+
-
*[[Melbourne/BBa_E0040|'''P1 5H 2''']]
+
-
Stored at -80
+
-
 
+
-
====Liquid Culture====
+
-
[[Melbourne/Growing up cells|Cultured]] the following in 5mL LB
+
-
*[[Melbourne/BBa_I15010|'''I15010 1''']] (Kan)
+
-
*[[Melbourne/BBa_I15010|'''I15010 2''']]
+
-
*[[Melbourne/BBa_R0084|'''P1 11H 1''']] (Amp)
+
-
*[[Melbourne/BBa_R0084|'''P1 11H 2''']]
+
-
 
+
-
Also placed Kan plate in the incubator to test antibiotic efficiency - no growth on 8/7
+
-
 
+
-
===7 July 2007===
+
-
 
+
-
*Made 10x TAE buffer
+
-
 
+
-
====Digest Gel====
+
-
*Prepared 8 lane 60mL [[Melbourne/Preparing an agarose gel|agarose gel]] with 1xTAE buffer.
+
-
*[[Melbourne/Loading a DNA gel|Loaded]] 20uL of digest samples from 6/7 in the following lane order.
+
-
*#[[Melbourne/BBa_J61035|'''P4 8J 1''']]
+
-
*#[[Melbourne/BBa_J61035|'''P4 8J 2''']]
+
-
*#[[Melbourne/BBa_E0040|'''P1 5H 1''']]
+
-
*#[[Melbourne/BBa_E0040|'''P1 5H 2''']]
+
-
*#[[Melbourne/BBa_E0241|'''P2 15L 1''']]
+
-
*#[[Melbourne/BBa_E0241|'''P2 15L 2''']]
+
-
 
+
-
====Glycerol Stocks====
+
-
The following [[Melbourne/Glycerol Stocks|glycerol stocks]] were made and dated 7/7:
+
-
*[[Melbourne/BBa_I15010|'''I15010 1''']]
+
-
*[[Melbourne/BBa_I15010|'''I15010 2''']]
+
-
*[[Melbourne/BBa_R0084|'''P1 11H 1''']]
+
-
*[[Melbourne/BBa_R0084|'''P1 11H 2''']]
+
-
Stored at -80
+
-
 
+
-
====Miniprep====
+
-
*Miniprepped the remains of the cultures and labelled with todays date 7/7.  Samples were eluted with TE buffer.
+
-
 
+
-
====Digest====
+
-
*[[Melbourne/Diagnostic Digest|Digested]] 5uL of each of the above miniprep DNA
+
-
**EcoRI/PstI in buffer 3
+
-
***[[Melbourne/BBa_I15010|'''I15010 1''']] (Kan)
+
-
***[[Melbourne/BBa_I15010|'''I15010 2''']]
+
-
***[[Melbourne/BBa_R0084|'''P1 11H 1''']] (Amp)
+
-
***[[Melbourne/BBa_R0084|'''P1 11H 2''']]
+
-
*Incubated for 3hours at 37degrees
+
-
*Added 5uL 6x loading dye and stored at -20
+
-
 
+
-
===8 July 2007===
+
-
 
+
-
====Transformation====
+
-
====Tranformation====
+
-
Resuspended and [[Melbourne/Transformation Protocol|transformed]] the following
+
-
*[[Melbourne/BBa_R0082|'''P1 15P''']](BBa_R0082, Omp R+, Amp)
+
-
*[[Melbourne/BBa_R0083|'''P1 17H''']](BBa_R0083, truncated BBa_R0082 Omp R+, Amp)
+
-
*[[Melbourne/BBa_E0430|'''P1 11A''']](BBa_E0430, EYFP(RBS+,LVA-,term) Amp)
+
-
*[[Melbourne/BBa_E0840|'''P1 16E''']](BBa_E0430; RBS,GFP,term; Amp)
+
-
The following was also retransformed due to colonies on previous plate appearing to be contaminants and failure of plasmid isolation from these colonies.
+
-
*[[Melbourne/BBa_Q04510|'''P2 13K''']] (BBa_Q04510, c1 inverter, Kan)
+
-
 
+
-
==Week 3==
+
-
===9 July 2007===
+
-
===10 July 2007===
+
-
===11 July 2007===
+
-
===12 July 2007===
+
-
===13 July 2007===
+
-
==Week 4==
+
-
===16 July 2007===
+
-
===17 July 2007===
+
-
===18 July 2007===
+
-
===19 July 2007===
+
-
===20 July 2007===
+
-
==Week 5==
+
-
===23 July 2007===
+
-
===24 July 2007===
+
-
===25 July 2007===
+
-
===26 July 2007===
+
-
===27 July 2007===
+
-
==Week 6==
+
-
===30 July 2007===
+
-
===31 July 2007===
+
-
===1  Aug 2007===
+
-
===2  Aug 2007===
+
-
===3  Aug 2007===
+

Latest revision as of 14:25, 26 October 2007

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Retrieved from "http://2007.igem.org/wiki/index.php/Melbourne/Lab_Notebook"