Team:NTU-Singapore/Wetlab

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='''Wet Lab Introduction'''=
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=Introduction=
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Wet laboratory experiments are held in our faculty laboratories and include the following experiments:<br>
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We employed the methodology used on basic engineering concepts: Abstraction, decoupling and standardization.  
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1) Creation of Standard Biobrick parts<br>
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2) Digestion and Ligation of NTU@iGEM System<br>
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3) Characterization of standard Biobrick promoters via GFP activity<br>
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4) System verification using OD measurement<br>
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<br>
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[[Image:wetlab_Introduction.jpg|924px x 580px|thumb|center|Our Wetlab Approach]]
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* An overview of how our experiments are conducted is represented in the above figure.<br>
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<br>
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Our major biobrick parts: Lysis protein generator, E7-Immunity protein generator, LsrA promoter were created using standard biobrick construction protocol. The genes for these parts were extracted from their wild bacteria strains (Escherichia Coli MG1655, BW and W3110) and modified to become standard biobrick modules. The parts were then verified by Gel Electrophoresis for its base pair length.
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<br><br>
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After the parts were created, our “system assembly factory” starts to fuse the individual biobrick parts to from devices, and eventually to our overall system. Again, Gel electrophoresis confirmed that our system assembly process was successful as the end products had base pair lengths that match our prediction.<br><br>
 +
Our Detection & Lysis system was tested with Ai2 induction and results have shown that our system works wonderfully!  On addition of Ai2 in LB growth culture, optical density measurement starts to show a constant value over time as compared to our control experiment which shows a trend of ever increasing OD values. This illustrates that when AI2 is added, cell transformed with our lysis systems starts producing lysis gene which cause cell lysis and death, and hence a constant OD values. Without induction of Ai2, cells continue to multiply, leading to higher OD values. 
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<br><br>
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While our system is being tested, we also performed characterization experiments for 2 promoters:<br>
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<br>
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• Existing LacI promoter which is regulated by lactose concentration<br>
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• NTU@iGEM new LsrA promoter which is regulated by Ai2 concentration.<br>
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<br>
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For more information characterization results, please see section: [https://2008.igem.org/Team:NTU-Singapore/Parts Characterization].<br>
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<br><br>
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<html>
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==Abstraction==
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var tags_before_clock = "<b>It is now "
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In abstraction we reduce the complexity of things by creating simple parts and developing these parts into higher complex devices and systems
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var tags_middle_clock = "on"
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var tags_after_clock  = "</b>"
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[[Image:NTU@iGEM_Abstraction_pic.JPG|900px]]
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==Decoupling==
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DaysofWeek = new Array()
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In decoupling, we decoupled each individual part from the others, devising them without consideration for the other existing part. Following which the fusion of different combination of components formed a working device
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  DaysofWeek[0]="Sunday"
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  DaysofWeek[1]="Monday"
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  DaysofWeek[3]="Wednesday"
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  DaysofWeek[4]="Thursday"
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  DaysofWeek[5]="Friday"
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  DaysofWeek[6]="Saturday"
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[[Image:NTU@iGEM Decoupling_pic.JPG|900px]]
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Months = new Array()
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  Months[0]="January"
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  Months[1]="February"
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  Months[2]="March"
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  Months[3]="April"
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  Months[4]="May"
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  Months[5]="June"
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  Months[6]="July"
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  Months[7]="August"
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  Months[8]="September"
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  Months[9]="October"
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  Months[11]="December"
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var date = dte.getDate()
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var month = Months[dte.getMonth()]
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var year = dte.getFullYear()
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==Standardization==
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var spc = " ";
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in standardization, we aim to establish standards that are widely used and agreed. We also allowed the predicament of components and compatibility of parts leading to a set of standards and protocols that could be used for future teams.
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Latest revision as of 05:46, 27 October 2008

Wet Lab Introduction

Wet laboratory experiments are held in our faculty laboratories and include the following experiments:

1) Creation of Standard Biobrick parts
2) Digestion and Ligation of NTU@iGEM System
3) Characterization of standard Biobrick promoters via GFP activity
4) System verification using OD measurement

Our Wetlab Approach
  • An overview of how our experiments are conducted is represented in the above figure.


Our major biobrick parts: Lysis protein generator, E7-Immunity protein generator, LsrA promoter were created using standard biobrick construction protocol. The genes for these parts were extracted from their wild bacteria strains (Escherichia Coli MG1655, BW and W3110) and modified to become standard biobrick modules. The parts were then verified by Gel Electrophoresis for its base pair length.

After the parts were created, our “system assembly factory” starts to fuse the individual biobrick parts to from devices, and eventually to our overall system. Again, Gel electrophoresis confirmed that our system assembly process was successful as the end products had base pair lengths that match our prediction.

Our Detection & Lysis system was tested with Ai2 induction and results have shown that our system works wonderfully! On addition of Ai2 in LB growth culture, optical density measurement starts to show a constant value over time as compared to our control experiment which shows a trend of ever increasing OD values. This illustrates that when AI2 is added, cell transformed with our lysis systems starts producing lysis gene which cause cell lysis and death, and hence a constant OD values. Without induction of Ai2, cells continue to multiply, leading to higher OD values.

While our system is being tested, we also performed characterization experiments for 2 promoters:

• Existing LacI promoter which is regulated by lactose concentration
• NTU@iGEM new LsrA promoter which is regulated by Ai2 concentration.

For more information characterization results, please see section: Characterization.


Retrieved from "http://2008.igem.org/Team:NTU-Singapore/Wetlab"