Team:Davidson-MissouriW/Notebook

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==Notebook==
==Notebook==
 +
'''May 25, 2010 to May 29, 2010'''
 +
<hr>
 +
<ul>
 +
<li>Davidson traveled to Missouri Western and further discussed project ideas</li>
 +
</ul>
 +
<br>
'''June 1, 2010'''
'''June 1, 2010'''
<hr>
<hr>
Line 9: Line 15:
<li>Verified RCBS with our sequences and researched codon optimization methods</li>
<li>Verified RCBS with our sequences and researched codon optimization methods</li>
<li>Analyzed probabilities for constructs A and B</li>
<li>Analyzed probabilities for constructs A and B</li>
 +
<li>Learned lab protocols and organized lab
 +
<li>Created list of needed parts and parts we need
</ul>  
</ul>  
<br>
<br>
Line 17: Line 25:
<li>Researched different ways to optimize a gene</li>
<li>Researched different ways to optimize a gene</li>
<li>Began simulation for Constructs A and B</li>
<li>Began simulation for Constructs A and B</li>
 +
<li>Learned miniprep and digestion protocols
 +
<li>Learned how to document gels
</ul>
</ul>
<br>
<br>
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<li>Researched restriction sites and created proposal for gene optimization</li>
<li>Researched restriction sites and created proposal for gene optimization</li>
<li>Continued work on simulation in Java and Matlab</li>
<li>Continued work on simulation in Java and Matlab</li>
-
 
+
<li>Learned about biobrick restriction sites and sticky ends
 +
<li>Ordered lox oligos
</ul>
</ul>
<br>
<br>
Line 35: Line 46:
<li>Analyzed the optimization of tetA sequence and researched properties of Lox sites
<li>Analyzed the optimization of tetA sequence and researched properties of Lox sites
<li>Debugged Simulations
<li>Debugged Simulations
 +
<li>Sent off parts for sequencing
 +
<li>Learned the transformation protocol
  </li>
  </li>
</ul>
</ul>
Line 44: Line 57:
<li>Began working to improve the Lancelator and created histograms for simulations
<li>Began working to improve the Lancelator and created histograms for simulations
  </li>
  </li>
 +
<li>Gel purified vector for lox ligations
 +
<li>Divided up constructs to be ligated
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Double digest on pBAD-RBS-RFP to make an RS fragment insert. </li>
+
<li>Double digest on pBAD-RBS-RFP to make an RS fragment insert </li>
<li>Worked on a Perl program that check for Bio Brick Restriction Sites
<li>Worked on a Perl program that check for Bio Brick Restriction Sites
 +
<li>Analyzed sequencing results using ApE program
 +
<li>BOUGHT HAWAIIAN SHIRTS!!!!!
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>EcoRI and PstI digest on pLac and pBad colonies.
+
<li>EcoRI and PstI digest on pLac and pBad colonies  
-
<li>Arabinose and IPTG concentration experiments started. </li>
+
<li>Arabinose and IPTG concentration experiments started </li>
<li>Continued work on the Lancelator program and began working on a different simulation for Construct C and D
<li>Continued work on the Lancelator program and began working on a different simulation for Construct C and D
 +
<li>Annealed lox oligos together
 +
<li>Gel purified pBad+RBS+Cre (“Cre construct” from 2010 igem kit plate)
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Picked 14 colonies from pLac-RBS-RFP and pLac-RBS-RFP-RBS plates.
+
<li>Picked 14 colonies from pLac-RBS-RFP and pLac-RBS-RFP-RBS plates
-
<li> Started inducer curve experiments.</li>
+
<li> Started inducer curve experiments</li>
<li>Linked Lancelator code to the website and continued simulations
<li>Linked Lancelator code to the website and continued simulations
 +
<li>Performed ligations involving lox sites
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Ligated S03736 into pSB1A7 vector.
+
<li>Ligated S03736 into pSB1A7 vector
-
<li>EcoRI/PstI digest on pLac-RBS-RFP-RBS colonies.</li>
+
<li>EcoRI/PstI digest on pLac-RBS-RFP-RBS colonies</li>
<li>Added new features to the Lancelator and worked on numerically representing Construct C for the Matlab simulation
<li>Added new features to the Lancelator and worked on numerically representing Construct C for the Matlab simulation
 +
<li>Performed colony PCR screening for the first ligations
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>EcoRI/PstI diagnostic digest performed on pBad-RBS-RFP colonies.  
+
<li>EcoRI/PstI diagnostic digest performed on pBad-RBS-RFP colonies   
-
<li>Inducer curve experiment started on pBad and pLac constructs. </li>
+
<li>Inducer curve experiment started on pBad and pLac constructs </li>
<li>Debugged Lancelator and finished numerical representation for the simulation
<li>Debugged Lancelator and finished numerical representation for the simulation
 +
<li>Size verified lox forward and lox reverse sites
 +
<li>Ligated “cre construct” into low, medium, and high copy kanamycin resistant vectors
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Ligated pLac-RBS-RFP + RBS-TetA.
+
<li>Ligated pLac-RBS-RFP + RBS-TetA
<li>IPTG experiment started on pLacI-RBS-RFP </li>
<li>IPTG experiment started on pLacI-RBS-RFP </li>
<li>Worked on the color schemes for the new Oligator
<li>Worked on the color schemes for the new Oligator
<li>Created a new construct that might be helpful for the biologists
<li>Created a new construct that might be helpful for the biologists
 +
<li>Planning for Amp+Tet experiments
 +
<li>First “cre construct” ligations failed
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>EcoRI/PstI diagnostic digest performed on pLac-RBS-RFP in pSB1A7 vector.
+
<li>EcoRI/PstI diagnostic digest performed on pLac-RBS-RFP in pSB1A7 vector
-
<li>IPTG experiment started on plates with pLac and pLacI constructs. </li>
+
<li>IPTG experiment started on plates with pLac and pLacI constructs </li>
<li>Began working on a Java code that will run simulations for all the constructs
<li>Began working on a Java code that will run simulations for all the constructs
<li>Researched on NP-Complete problems
<li>Researched on NP-Complete problems
 +
<li>Religated “cre construct”
 +
<li>Lox sites sent off for sequencing
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Flourimeter data collected from IPTG experiment.
+
<li>Flourimeter data collected from IPTG experiment
-
<li>Ligated pLacI-RBS-RFP + RBS-TetA. </li>
+
<li>Ligated pLacI-RBS-RFP + RBS-TetA </li>
<li>Worked on graphics of the Java program
<li>Worked on graphics of the Java program
<li>Studied specific NP-Complete problems and brainstormed ways to incorporate them in our project
<li>Studied specific NP-Complete problems and brainstormed ways to incorporate them in our project
 +
<li>Lox sites were frozen down and entered in GCAT-alog
 +
<li>Agar plates were prepared for Amp+Tet experiments
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Picked 4 colonies from pLacI-RBS-RFP-RBS-TetA plates.
+
<li>Picked 4 colonies from pLacI-RBS-RFP-RBS-TetA plates  
-
<li>IPTG experiment started using newly created IPTG. </li>
+
<li>IPTG experiment started using newly created IPTG  </li>
<li>Debugged the Java program  
<li>Debugged the Java program  
<li>Created a new page for the GCAT server
<li>Created a new page for the GCAT server
 +
<li>Planned kanamycin experiments
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Digested TetA with EcoRI/NheI.
+
<li>Digested TetA with EcoRI/NheI
-
<li>Annealed oligos for optimized and deoptimized TetA segment 1. </li>
+
<li>Annealed oligos for optimized and deoptimized TetA segment 1  </li>
<li>Improved the Java program
<li>Improved the Java program
<li>Began categorizing the knapsack problem into subparts
<li>Began categorizing the knapsack problem into subparts
 +
<li>Successfully ligated ptet+different lox sites
 +
<li>Planned different combinations of variant lox sites
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Ligated annealed TetA oligos into digested TetA.
+
<li>Ligated annealed TetA oligos into digested TetA
-
<li>Inducer curve experiment started on pLac and pLacI constructs. </li>
+
<li>Inducer curve experiment started on pLac and pLacI constructs  </li>
<li>Added buttons to the java program
<li>Added buttons to the java program
<li>Brainstormed biological implications in different NP-Complete problems
<li>Brainstormed biological implications in different NP-Complete problems
 +
<li>Confirmed successful ligations of lox sites from sequencing results
 +
<li>First pilot test for Amp+Tet experiments
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Preparative NheI/BamHI digest performed on TetA.
+
<li>Preparative NheI/BamHI digest performed on TetA
-
<li>Preparative EcoRI/NheI digest performed on TetA. </li>
+
<li>Preparative EcoRI/NheI digest performed on TetA </li>
<li>Restructuring and reorganizing the Java program
<li>Restructuring and reorganizing the Java program
<li>Reviewed numerical algorithms and its relation to NP-Complete problems
<li>Reviewed numerical algorithms and its relation to NP-Complete problems
 +
<li>Researched data on possible transcription terminator in Tet A
 +
<li>Failure of cre experiments
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Annealed optimized and deoptimized segment 1 TetA ligated into vector.
+
<li>Annealed optimized and deoptimized segment 1 TetA ligated into vector  
-
<li>Annealed optimized and deoptimized segment 2 TetA ligatied into vector.</li>
+
<li>Annealed optimized and deoptimized segment 2 TetA ligatied into vector</li>
<li>Added weight characteristic to the fluorescent proteins in the Java code
<li>Added weight characteristic to the fluorescent proteins in the Java code
<li>Continued exploration of NP-Complete problems
<li>Continued exploration of NP-Complete problems
 +
<li>Ligations to construct tet constructs
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Inducer curve experiment started on pLac and pLacI constructs.
+
<li>Inducer curve experiment started on pLac and pLacI constructs  
-
<li>Picked colonies from TetA segment 1 & 2 optimized & deoptimized plates. </li>
+
<li>Picked colonies from TetA segment 1 & 2 optimized & deoptimized plates </li>
<li>Added histograms to the Java Code
<li>Added histograms to the Java Code
<li>Researched and analyzed the effectiveness of Cre
<li>Researched and analyzed the effectiveness of Cre
 +
<li>Size verification of ptet+variant lox sites
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul><li>Added the option for the user to input the weights of the various modules and began work on custom construct builder
<ul><li>Added the option for the user to input the weights of the various modules and began work on custom construct builder
-
<li>Picked colonies from TetA segment 1 & 2 optimized & deoptimized plates. </li>
+
<li>Picked colonies from TetA segment 1 & 2 optimized & deoptimized plates </li>
 +
<li>Planned to ligate pBad+RBS+Cre in ampicillin resistant vectors
 +
<li>Researched plasmid partitioning
</ul>
</ul>
<br>
<br>
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<li>Explored the set covering problem
<li>Explored the set covering problem
<li>Researched applications of the knapsack problem in cryptography
<li>Researched applications of the knapsack problem in cryptography
 +
<li>Ligated ptet+LoxP+RBS+RFP
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Tet titration experiment performed on pLac and pLacI +RBS-RFP-RBS-TetA.
+
<li>Tet titration experiment performed on pLac and pLacI +RBS-RFP-RBS-TetA
-
<li>Ligated TetA onto pLac and pLacI +RBS-RFP-RBS-TetA with no terminator. </li>
+
<li>Ligated TetA onto pLac and pLacI +RBS-RFP-RBS-TetA with no terminator  </li>
<li>Some final work on the simulation program
<li>Some final work on the simulation program
<li>Discussed new ideas such as integer programming
<li>Discussed new ideas such as integer programming
 +
<li>PCR screening of “RFP construct”
 +
<li>Ptet+variant lox sites were frozen down
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Preparative BamHI/NheI digest performed on TetA.
+
<li>Preparative BamHI/NheI digest performed on TetA  
-
<li>New experiment protocol developed for reduced vector background noise.   </li>
+
<li>New experiment protocol developed for reduced vector background noise  </li>
<li>Added terminators to the program
<li>Added terminators to the program
<li>Ran test trials of the program and analyzed its results
<li>Ran test trials of the program and analyzed its results
 +
<li>Planned experiments for using Cre as “front” and “back” vector
 +
<li>Ligated first “floxed” constructs
</ul>
</ul>
<br>
<br>
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<li>Continued work on the categorization of the knapsack problem
<li>Continued work on the categorization of the knapsack problem
<li>Researched Instant Insanity problem
<li>Researched Instant Insanity problem
 +
<li>PCR screening for “floxed constructs”
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Ligated S04446 and S04447 to J31007.
+
<li>Ligated S04446 and S04447 to J31007
-
<li>Tet titration experiment performed on pLac and pLacI +RBS-RFP-RBS-TetA. </li>
+
<li>Tet titration experiment performed on pLac and pLacI +RBS-RFP-RBS-TetA </li>
<li>Added direction of terminators in the simulation
<li>Added direction of terminators in the simulation
<li>Researched cryptography application
<li>Researched cryptography application
 +
<li>Size verified “floxed constructs”
 +
<li>Digested “Cre construct” as front and back vector
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Ligated S04448 to RBS-TetA.
+
<li>Ligated S04448 to RBS-TetA
-
<li>Ligated S04449 to RBS-TetA.</li>
+
<li>Ligated S04449 to RBS-TetA</li>
<li>Fixed memory leak in the simulation program
<li>Fixed memory leak in the simulation program
<li>Researched integer programming
<li>Researched integer programming
 +
<li>Sent off “floxed constructs" for sequencing
 +
<li>Ligated “floxed constructs” into pBad+RBS+Cre as “front” and “back” vector 
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>IPTG experiment started using pre-induced method.  
+
<li>IPTG experiment started using pre-induced method   
-
<li>IPTG experiment started using varying levels of tetracycline. </li>
+
<li>IPTG experiment started using varying levels of tetracycline  </li>
<li>Worked on animation for the simulation and continued analysis of the results of the simulation
<li>Worked on animation for the simulation and continued analysis of the results of the simulation
<li>Discussed integer programming as a group
<li>Discussed integer programming as a group
 +
<li>Data suggests something wrong with “cre construct”
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Picked optimized and deoptimized TetA colonies.
+
<li>Picked optimized and deoptimized TetA colonies
-
<li>Flourimeter data collected from IPTG experiment. </li>
+
<li>Flourimeter data collected from IPTG experiment  </li>
<li>Revised some features on the program
<li>Revised some features on the program
<li>Began working on theoretical probability models of the different constructs
<li>Began working on theoretical probability models of the different constructs
 +
<li>Further experiments with “cre construct”
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Used RFLP to screen for candidate TetA clones.
+
<li>Used RFLP to screen for candidate TetA clones  
-
<li>Transformed various pLac and pLacI constructs into MG1655 cells. </li>
+
<li>Transformed various pLac and pLacI constructs into MG1655 cells </li>
 +
<li>Experiments suggest pBad+RBS+Cre from iGem 2010 kit plate is incorrect
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Used RFLP to screen for candidate TetA clones.
+
<li>Used RFLP to screen for candidate TetA clones  
-
<li>Transformed various pLac and pLacI constructs into MG1655 cells. </li>
+
<li>Transformed various pLac and pLacI constructs into MG1655 cells </li>
<li>Added slider feature to the simulation program
<li>Added slider feature to the simulation program
<li>Continued work on the theoretical probability model for Construct A of size 3
<li>Continued work on the theoretical probability model for Construct A of size 3
 +
<li>Tested “cre construct” from 2009 iGem kit plate
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Used RFLP to screen for candidate TetA clones.
+
<li>Used RFLP to screen for candidate TetA clones  
-
<li>EcoRI/PstI diagnostic digest performed on K199150. </li>
+
<li>EcoRI/PstI diagnostic digest performed on K199150 </li>
<li>Added custom prefixes and suffixes to the Oligator
<li>Added custom prefixes and suffixes to the Oligator
<li>Found a general equation for finding theoretical probabilities for Construct A of size 3
<li>Found a general equation for finding theoretical probabilities for Construct A of size 3
 +
<li>Sent off “floxed sites for sequencing”
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Ligated segment 2 optimized and deoptimzed TetA behind segment 1.
+
<li>Ligated segment 2 optimized and deoptimzed TetA behind segment 1
-
<li>IPTG experiment started on I715039-1 and I715039-2. </li>
+
<li>IPTG experiment started on I715039-1 and I715039-2 </li>
<li>Began analyzing MW optimization code
<li>Began analyzing MW optimization code
<li>Finished theoretical probabilities for Construct A of size 3
<li>Finished theoretical probabilities for Construct A of size 3
 +
<li>Cre construct from 2009 plate was incorrect as well
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Cell count data collected from IPTG experiments.
+
<li>Cell count data collected from IPTG experiments
-
<li>Flourimeter data collected from IPTG experiments.  </li>
+
<li>Flourimeter data collected from IPTG experiments </li>
<li>Worked on a separate program using the CAI equation
<li>Worked on a separate program using the CAI equation
<li>Began working on theoretical probabilities for Construct A of size 4
<li>Began working on theoretical probabilities for Construct A of size 4
 +
<li>Ligated more “floxed constructs”
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Tet titration experiment started using pSB3T5 vector.
+
<li>Tet titration experiment started using pSB3T5 vector
-
<li>Tet titration experiment started using optimized and deoptimized TetA. </li>
+
<li>Tet titration experiment started using optimized and deoptimized TetA </li>
<li>Added changes to the optimizer code
<li>Added changes to the optimizer code
<li>Researched how to solve a system of recursive relations into closed form
<li>Researched how to solve a system of recursive relations into closed form
 +
<li>Decided to ligate cre into pBad+RBS and pLac+RBS
</ul>
</ul>
<br>
<br>
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<hr>
<hr>
<ul>
<ul>
-
<li>Cell count data collected from Tet titrations.
+
<li>Cell count data collected from Tet titrations
-
<li>Tet titration experiment started using optimized and deoptimized TetA. </li>
+
<li>Tet titration experiment started using optimized and deoptimized TetA </li>
-
</ul>
+
<li>Fixed the error in the CAI optimization program
-
<br>
+
<li>Continued analyzing systems of recursive relations
-
'''June 3, 2010'''
+
<li>Ligated pBad+RBS+Cre and pLac+RBS+Cre
-
<hr>
+
-
<ul>
+
-
<li>Mini prepped, diagnostic RP digest, preparative digest, ligation, pBAD+RBS-RFP </li>
+
</ul>
</ul>
<br>
<br>
Line 312: Line 367:
<hr>
<hr>
<ul>
<ul>
-
<li>Flourimeter data collected on IPTG experiment. </li>
+
<li>Flourimeter data collected on IPTG experiment </li>
 +
<li>Adapted the Oligator page for the Optimization program, now called the Optimus
 +
<li>Made progress on solving the system of recursive relations that we have
 +
<li>Performed IPTG and tet experiments
 +
<li>PCR screened both “cre constructs”
 +
<li>pBad+RBS+Cre ligation unsuccesful
</ul>
</ul>
<br>
<br>
Line 318: Line 378:
<hr>
<hr>
<ul>
<ul>
-
<li>IPTG experiment started on optimized and deoptmized TetA constructs.
+
<li>IPTG experiment started on optimized and deoptmized TetA constructs
-
<li>Passage I715039-1 and I715039-2 experiments started. </li>
+
<li>Passage I715039-1 and I715039-2 experiments started  </li>
 +
<li>Started to link the ruby program on the webpage
 +
<li>Finished theoretical probabilities for Construct A of size 4
 +
<li>"Floxed constructs" ligated into pLac+RBS+Cre
</ul>
</ul>
<br>'''July 22, 2010'''
<br>'''July 22, 2010'''
<hr>
<hr>
<ul>
<ul>
-
<li>IPTG experiment started using varying tetracycline concentrations.
+
<li>IPTG experiment started using varying tetracycline concentrations
-
<li>Cell count data collected on IPTG TetA experiment. </li>
+
<li>Cell count data collected on IPTG TetA experiment </li>
 +
<li>Linked the Optimus on the webpage
 +
<li>Started theoretical probabilities for Construct A of size 5
 +
<li>Cre was religated into pBad+RBS
</ul>
</ul>
<br>
<br>
Line 331: Line 397:
<hr>
<hr>
<ul>
<ul>
-
<li>Cell count data collected from IPTG experiment..
+
<li>Cell count data collected from IPTG experiment
-
<li>Flourimeter data collected from IPTG experiment. </li>
+
<li>Flourimeter data collected from IPTG experiment </li>
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<li>Worked on the tutorial for the Optimus and added various features
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<li>Worked on a Matlab tool based on the pancake problem that can find the theoretical probabilities of different modules in different positions
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<li>Last of the “floxed constructs” were ligated
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Latest revision as of 04:33, 26 October 2010

Notebook

May 25, 2010 to May 29, 2010

  • Davidson traveled to Missouri Western and further discussed project ideas


June 1, 2010

  • Tested to find lethal concentration of Tet in plated and liquid mediums
  • Verified RCBS with our sequences and researched codon optimization methods
  • Analyzed probabilities for constructs A and B
  • Learned lab protocols and organized lab
  • Created list of needed parts and parts we need


June 2, 2010

  • Mini prepped, diagnostic RP digest, preparative digest, ligation, pBAD+RBS-RFP
  • Researched different ways to optimize a gene
  • Began simulation for Constructs A and B
  • Learned miniprep and digestion protocols
  • Learned how to document gels


June 3, 2010

  • Mini prepped, diagnostic RP digest, preparative digest, ligation, pBAD+RBS-RFP
  • Researched restriction sites and created proposal for gene optimization
  • Continued work on simulation in Java and Matlab
  • Learned about biobrick restriction sites and sticky ends
  • Ordered lox oligos


June 4, 2010

  • Mini prepped RBS-TetA-TT
  • RX Fragment digestion for RBS-TetA-TT
  • Analyzed the optimization of tetA sequence and researched properties of Lox sites
  • Debugged Simulations
  • Sent off parts for sequencing
  • Learned the transformation protocol


June 7, 2010

  • Ligate and plasmid check for pSB1A2 or pSB1AK3
  • Began working to improve the Lancelator and created histograms for simulations
  • Gel purified vector for lox ligations
  • Divided up constructs to be ligated


June 8, 2010

  • Double digest on pBAD-RBS-RFP to make an RS fragment insert
  • Worked on a Perl program that check for Bio Brick Restriction Sites
  • Analyzed sequencing results using ApE program
  • BOUGHT HAWAIIAN SHIRTS!!!!!


June 9, 2010

  • EcoRI and PstI digest on pLac and pBad colonies
  • Arabinose and IPTG concentration experiments started
  • Continued work on the Lancelator program and began working on a different simulation for Construct C and D
  • Annealed lox oligos together
  • Gel purified pBad+RBS+Cre (“Cre construct” from 2010 igem kit plate)


June 10, 2010

  • Picked 14 colonies from pLac-RBS-RFP and pLac-RBS-RFP-RBS plates
  • Started inducer curve experiments
  • Linked Lancelator code to the website and continued simulations
  • Performed ligations involving lox sites


June 11, 2010

  • Ligated S03736 into pSB1A7 vector
  • EcoRI/PstI digest on pLac-RBS-RFP-RBS colonies
  • Added new features to the Lancelator and worked on numerically representing Construct C for the Matlab simulation
  • Performed colony PCR screening for the first ligations


June 14, 2010

  • EcoRI/PstI diagnostic digest performed on pBad-RBS-RFP colonies
  • Inducer curve experiment started on pBad and pLac constructs
  • Debugged Lancelator and finished numerical representation for the simulation
  • Size verified lox forward and lox reverse sites
  • Ligated “cre construct” into low, medium, and high copy kanamycin resistant vectors


June 15, 2010

  • Ligated pLac-RBS-RFP + RBS-TetA
  • IPTG experiment started on pLacI-RBS-RFP
  • Worked on the color schemes for the new Oligator
  • Created a new construct that might be helpful for the biologists
  • Planning for Amp+Tet experiments
  • First “cre construct” ligations failed


June 16, 2010

  • EcoRI/PstI diagnostic digest performed on pLac-RBS-RFP in pSB1A7 vector
  • IPTG experiment started on plates with pLac and pLacI constructs
  • Began working on a Java code that will run simulations for all the constructs
  • Researched on NP-Complete problems
  • Religated “cre construct”
  • Lox sites sent off for sequencing


June 17, 2010

  • Flourimeter data collected from IPTG experiment
  • Ligated pLacI-RBS-RFP + RBS-TetA
  • Worked on graphics of the Java program
  • Studied specific NP-Complete problems and brainstormed ways to incorporate them in our project
  • Lox sites were frozen down and entered in GCAT-alog
  • Agar plates were prepared for Amp+Tet experiments


June 18, 2010

  • Picked 4 colonies from pLacI-RBS-RFP-RBS-TetA plates
  • IPTG experiment started using newly created IPTG
  • Debugged the Java program
  • Created a new page for the GCAT server
  • Planned kanamycin experiments


June 21, 2010

  • Digested TetA with EcoRI/NheI
  • Annealed oligos for optimized and deoptimized TetA segment 1
  • Improved the Java program
  • Began categorizing the knapsack problem into subparts
  • Successfully ligated ptet+different lox sites
  • Planned different combinations of variant lox sites


June 22, 2010

  • Ligated annealed TetA oligos into digested TetA
  • Inducer curve experiment started on pLac and pLacI constructs
  • Added buttons to the java program
  • Brainstormed biological implications in different NP-Complete problems
  • Confirmed successful ligations of lox sites from sequencing results
  • First pilot test for Amp+Tet experiments


June 23, 2010

  • Preparative NheI/BamHI digest performed on TetA
  • Preparative EcoRI/NheI digest performed on TetA
  • Restructuring and reorganizing the Java program
  • Reviewed numerical algorithms and its relation to NP-Complete problems
  • Researched data on possible transcription terminator in Tet A
  • Failure of cre experiments


June 24, 2010

  • Annealed optimized and deoptimized segment 1 TetA ligated into vector
  • Annealed optimized and deoptimized segment 2 TetA ligatied into vector
  • Added weight characteristic to the fluorescent proteins in the Java code
  • Continued exploration of NP-Complete problems
  • Ligations to construct tet constructs


June 27, 2010

  • Inducer curve experiment started on pLac and pLacI constructs
  • Picked colonies from TetA segment 1 & 2 optimized & deoptimized plates
  • Added histograms to the Java Code
  • Researched and analyzed the effectiveness of Cre
  • Size verification of ptet+variant lox sites


June 28, 2010

  • Added the option for the user to input the weights of the various modules and began work on custom construct builder
  • Picked colonies from TetA segment 1 & 2 optimized & deoptimized plates
  • Planned to ligate pBad+RBS+Cre in ampicillin resistant vectors
  • Researched plasmid partitioning


June 29, 2010

  • Screened segment 1 optimized and deoptimized candidate clones using RFLP
  • Screened segment 2 optimized and deoptimized candidate clones using RFLP
  • Finished the simulation program and exported it
  • Explored the set covering problem
  • Researched applications of the knapsack problem in cryptography
  • Ligated ptet+LoxP+RBS+RFP


June 30, 2010

  • Tet titration experiment performed on pLac and pLacI +RBS-RFP-RBS-TetA
  • Ligated TetA onto pLac and pLacI +RBS-RFP-RBS-TetA with no terminator
  • Some final work on the simulation program
  • Discussed new ideas such as integer programming
  • PCR screening of “RFP construct”
  • Ptet+variant lox sites were frozen down


July 1, 2010

  • Preparative BamHI/NheI digest performed on TetA
  • New experiment protocol developed for reduced vector background noise
  • Added terminators to the program
  • Ran test trials of the program and analyzed its results
  • Planned experiments for using Cre as “front” and “back” vector
  • Ligated first “floxed” constructs


July 2, 2010

  • Mini prepped, diagnostic RP digest, preparative digest, ligation, pBAD+RBS-RFP
  • Continued work on the categorization of the knapsack problem
  • Researched Instant Insanity problem
  • PCR screening for “floxed constructs”


July 6, 2010

  • Ligated S04446 and S04447 to J31007
  • Tet titration experiment performed on pLac and pLacI +RBS-RFP-RBS-TetA
  • Added direction of terminators in the simulation
  • Researched cryptography application
  • Size verified “floxed constructs”
  • Digested “Cre construct” as front and back vector


July 7, 2010

  • Ligated S04448 to RBS-TetA
  • Ligated S04449 to RBS-TetA
  • Fixed memory leak in the simulation program
  • Researched integer programming
  • Sent off “floxed constructs" for sequencing
  • Ligated “floxed constructs” into pBad+RBS+Cre as “front” and “back” vector


July 8, 2010

  • IPTG experiment started using pre-induced method
  • IPTG experiment started using varying levels of tetracycline
  • Worked on animation for the simulation and continued analysis of the results of the simulation
  • Discussed integer programming as a group
  • Data suggests something wrong with “cre construct”


July 9, 2010

  • Picked optimized and deoptimized TetA colonies
  • Flourimeter data collected from IPTG experiment
  • Revised some features on the program
  • Began working on theoretical probability models of the different constructs
  • Further experiments with “cre construct”


July 10, 2010

  • Used RFLP to screen for candidate TetA clones
  • Transformed various pLac and pLacI constructs into MG1655 cells
  • Experiments suggest pBad+RBS+Cre from iGem 2010 kit plate is incorrect


July 12, 2010

  • Used RFLP to screen for candidate TetA clones
  • Transformed various pLac and pLacI constructs into MG1655 cells
  • Added slider feature to the simulation program
  • Continued work on the theoretical probability model for Construct A of size 3
  • Tested “cre construct” from 2009 iGem kit plate


July 13, 2010

  • Used RFLP to screen for candidate TetA clones
  • EcoRI/PstI diagnostic digest performed on K199150
  • Added custom prefixes and suffixes to the Oligator
  • Found a general equation for finding theoretical probabilities for Construct A of size 3
  • Sent off “floxed sites for sequencing”


July 14, 2010

  • Ligated segment 2 optimized and deoptimzed TetA behind segment 1
  • IPTG experiment started on I715039-1 and I715039-2
  • Began analyzing MW optimization code
  • Finished theoretical probabilities for Construct A of size 3
  • Cre construct from 2009 plate was incorrect as well


July 15, 2010

  • Cell count data collected from IPTG experiments
  • Flourimeter data collected from IPTG experiments
  • Worked on a separate program using the CAI equation
  • Began working on theoretical probabilities for Construct A of size 4
  • Ligated more “floxed constructs”


July 16, 2010

  • Tet titration experiment started using pSB3T5 vector
  • Tet titration experiment started using optimized and deoptimized TetA
  • Added changes to the optimizer code
  • Researched how to solve a system of recursive relations into closed form
  • Decided to ligate cre into pBad+RBS and pLac+RBS


July 19, 2010

  • Cell count data collected from Tet titrations
  • Tet titration experiment started using optimized and deoptimized TetA
  • Fixed the error in the CAI optimization program
  • Continued analyzing systems of recursive relations
  • Ligated pBad+RBS+Cre and pLac+RBS+Cre


July 20, 2010

  • Flourimeter data collected on IPTG experiment
  • Adapted the Oligator page for the Optimization program, now called the Optimus
  • Made progress on solving the system of recursive relations that we have
  • Performed IPTG and tet experiments
  • PCR screened both “cre constructs”
  • pBad+RBS+Cre ligation unsuccesful


July 21, 2010

  • IPTG experiment started on optimized and deoptmized TetA constructs
  • Passage I715039-1 and I715039-2 experiments started
  • Started to link the ruby program on the webpage
  • Finished theoretical probabilities for Construct A of size 4
  • "Floxed constructs" ligated into pLac+RBS+Cre


July 22, 2010

  • IPTG experiment started using varying tetracycline concentrations
  • Cell count data collected on IPTG TetA experiment
  • Linked the Optimus on the webpage
  • Started theoretical probabilities for Construct A of size 5
  • Cre was religated into pBad+RBS



July 23, 2010

  • Cell count data collected from IPTG experiment
  • Flourimeter data collected from IPTG experiment
  • Worked on the tutorial for the Optimus and added various features
  • Worked on a Matlab tool based on the pancake problem that can find the theoretical probabilities of different modules in different positions
  • Last of the “floxed constructs” were ligated



July 26, 2010

  • Missouri traveled to Davidson to work on wiki and presentation materials
  • The two campuses reunite and share recent findings



July 27, 2010

  • Began work on the team wiki



October 25, 2010

  • Finalizing team wiki


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