Team:Johns Hopkins/Notebook

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Notebook

Contents

Files

iGEM Groups 1.0
iGEM Groups 2.01

Important reminders and notes

 [Can make general comments here, so they don't get lost in peoples e-mail boxes]
[For anyone not part of the iGEM team who would like to see the gels we're running: 1. Click the links provided 2. login: bluejays
July 11: Primers for group 1 were delivered yesterday
July 11: Lab meeting at 7:30PM in the lab to go over miniprep protocol
Tuesday July 15: Lab meeting at 6:30PM with Jessica. Have status reports ready. Bring labtop if you can

Status Reports

The status reports of each group below will continuously be updated as we work on the biobricks. The following PDFs contain progressive versions of our status reports as we continue through the sex detector project; they are added weekly. To learn more about each biobrick, please refer to the Biobrick page.

Status Report 2.1 - 07/12/08

GROUP 1: Fluorescent Proteins

 status report by: Ingrid (work done by James)
 Part no.: BBa_K110017
 Part Description: yESapphire RtL
 Part Location (in build a genome lab): In James and Jasper's PCR product Box,
 Stainless Steel 4 degree
 PCR successful?; Yes
 Cloning of PCR product successful: Y/N
 Sequencing of cloned PCR product successful: Not done
 Joining of validated part to adjacent part(s) status: Not done
 Problems to be solved: The PCR of this part yielded a very large product
 Current status of this part:
 status report by: Ingrid (work done by James)
 Part no.: BBa_K110010
 Part Description: yESapphire LtR
 Part Location (in build a genome lab): In James and Jasper's PCR product Box, 
 Stainless Steel 4 degree
 PCR successful?; Yes
 Cloning of PCR product successful: Not done 
 Sequencing of cloned PCR product successful: No
 Joining of validated part to adjacent part(s) status: Not done
 Problems to be solved: The PCR of this part yielded a very large product
 Current status of this part:
 status report by: Ingrid 
 Part no.: BBa_K110018 
 Part Description: mCherry: 3 to 5
 Part Location (in build a genome lab): In silver fridge by door
 PCR successful?; Yes 
 (http://baderlab.bme.jhu.edu:8888/moodle18/mod/data/view.php?d=4&rid=1471)
 Cloning of PCR product successful: Not done 
 Sequencing of cloned PCR product successful: No
 Joining of validated part to adjacent part(s) status: Not done
 Problems to be solved: Had some extra unknown products, with unknown bands in the gel
 Current status of this part: Done with touchdown PCR and gel. Next step is to clone product.
 status report by: Ingrid 
 Part no.: BBa_K110019
 Part Description: mCherry: 5 to 3
 Part Location (in build a genome lab): In silver fridge by door
 PCR successful?; Yes 
 (http://baderlab.bme.jhu.edu:8888/moodle18/mod/data/view.php?d=4&rid=1471)
 Cloning of PCR product successful: Not done 
 Sequencing of cloned PCR product successful: No
 Joining of validated part to adjacent part(s) status: Not done
 Problems to be solved: Had some extra unknown products, with unknown bands in the gel
 Current status of this part: Done with touchdown PCR and gel. Next step is to clone product.
 status report by: Ingrid 
 Part no.: BBa_K110020 
 Part Description: Venus Enhanced YFP
 Part Location (in build a genome lab): In silver fridge by door
 PCR successful?; Yes 
 (http://baderlab.bme.jhu.edu:8888/moodle18/mod/data/view.php?d=4&rid=1471)
 Cloning of PCR product successful: Not done 
 Sequencing of cloned PCR product successful: No
 Joining of validated part to adjacent part(s) status: Not done
 Problems to be solved: Had some extra unknown products, with unknown bands in the gel
 Current status of this part: Done with touchdown PCR and gel. Next step is to clone product.
 status report by: Ingrid 
 Part no.: BBa_K110021 
 Part Description: Venus Enhanced YFP
 Part Location (in build a genome lab): In silver fridge by door
 PCR successful?; Yes 
 (http://baderlab.bme.jhu.edu:8888/moodle18/mod/data/view.php?d=4&rid=1471)
 Cloning of PCR product successful: Not done 
 Sequencing of cloned PCR product successful: No
 Joining of validated part to adjacent part(s) status: Not done
 Problems to be solved: Had some extra unknown products, with unknown bands in the gel
 Current status of this part: Done with touchdown PCR and gel. Next step is to clone product.

GROUP 2: MATa Specific-promoters

 Status report by Allison and Nate
 Part no.: BBa_K110008
 Part Description: MFA1 (L+R)
 Part Location: in a labeled box, second shelf from the top, -20
 degrees C refrigerator next to front door
 Date: 7/10/08
 PCR successful? Yes
 http://baderlab.bme.jhu.edu:8888/moodle18/mod/data/view.php?d=4&advanced=0&paging=&page=33
 Cloning of PCR product successful: in progress
 Sequencing of cloned PCR product successful: not done
 Joining of validated part to adjacent part(s) status: not done
 Problems to be solved: to be determined
 Current status of this part: PCR was being troubleshooted, appeared to
 have good results with regular PCR protocol (not touchdown) in which
 there was a constant annealing temperature of 55 degrees C - see gel
 Status report by Allison and Nate
 Part no.: BBa_K110016
 Part Description: Ste2 (R+L)
 Part Location: in a labeled box, second shelf from the top, -20
 degrees C refrigerator next to front door
 Date: 7/10/08
 PCR successful? Yes
 http://baderlab.bme.jhu.edu:8888/moodle18/mod/data/view.php?d=4&advanced=0&paging=&page=33
 Cloning of PCR product successful: in progress
 Sequencing of cloned PCR product successful: not done
 Joining of validated part to adjacent part(s) status: not done
 Problems to be solved: to be determined
 Current status of this part: Both PCR protocols (touchdown and second
 PCR with constant annealing temperature) produced product of the
 correct size. BBa_K110016 was used as a control in the second PCR with
 BBa_K110008.
 Status report by Allison and Nate
 Part no.: BBa_K110008
 Part Description: MFA1 (L+R)
 Part Location: same as above, plates are at 4 degrees refrigerator near front door
 Date: 7/14/08
 PCR successful? Yes
 Cloning of PCR product of successful? There were mainly light blue colonies 
  (only a couple white colonies)
 Sequencing of cloned PCR product successful: not done
 Joining of validated part to adjacent part(s) status: not done
 Problems to be solved: Ligation
 Current status of this part: plates are at 4 degrees; another ligation/transformation 
  will be completed soon
 Status report by Allison and Nate
 Part no.: BBa_K110016
 Part Description: Ste2 (R+L)
 Part Location: in a labeled box, second shelf from the top, -20 degrees C refrigerator next to 
  front door; plates at 4 degrees
 Date: 7/14/08
 PCR successful? Yes
 Cloning of PCR product successful: There were many blue colonies (similar to the plate of BB_K110008)
 Sequencing of cloned PCR product successful: not done
 Joining of validated part to adjacent part(s) status: not done
 Problems to be solved: Ligation
 Current status of this part: plates are at 4 degrees; another ligation/transformation 
  will be completed soon

GROUP 3: Short two way stops

 Date: 7/11/08
 status report by: James
 Part no.: BBa_K110011
 Part Description: Between-bud 27-W FRS2-C LtR
 Part Location (in build a genome lab): In James and Jasper's PCR product Box,
  Stainless Steel 4 degree
 PCR successful?; Yes
 Cloning of PCR product successful: Yes (will come soon; I can put it 
  in the wiki to make it easier for you)
 Sequencing of cloned PCR product successful: No
 Joining of validated part to adjacent part(s) status: Not done
 Problems to be solved:
 Current status of this part:  Miniprep of Overnight 
  cultures will be completed today
 status report by: James
 Part no.: BBa_K110012
 Part Description: Between STE2-W and BST1-C LtR
 Part Location (in build a genome lab): In James and Jasper's PCR product Box, 
  Stainless Steel 4 degree
 PCR successful?; Yes
 Cloning of PCR product successful: Yes (will come soon; I can put it in the 
  wiki to make it easier for you)
 Sequencing of cloned PCR product successful: No
 Joining of validated part to adjacent part(s) status: Not done
 Problems to be solved:
 Current status of this part:  Miniprep of Overnight cultures will be completed today
 status report by: James
 Part no.: BBa_K110013
 Part Description: Between-SWP82-W and EMP47-C LtR
 Part Location (in build a genome lab): In James and Jasper's PCR product Box,
  Stainless Steel 4 degree
 PCR successful?; No
 Cloning of PCR product successful: No (will come soon; I can put it in the wiki 
  to make it easier for you)
 Sequencing of cloned PCR product successful: No
 Joining of validated part to adjacent part(s) status: Not done
 Problems to be solved: The PCR of this part yielded a very large product
 Current status of this part:

GROUP 4: Long Two-way Stops & Mat(alpha) specific promotors

 [Jaime- Please split]
 Date: 7/10/08
 Status report by: Jaime Liu
 Part no.: BBa_K110001, BBa_K110003, BBa_K110005, BBa_K110006
 Part Description:
 BBa_K110001 - Between-bud 27-W FRS2-C + 200bp into each gene LtR
 BBa_K110003 - Between-SWP82-W and EMP47-C +200 into each gene LtR
 BBa_K110005 - MFalpha2 LtR
 BBa_K110006 - MFalpha1 LtR
 Part Location (in build a genome lab): In 4C fridge #2
 PCR successful?; Y/N (link such as this)- Yes
 BBa_K110001, BBa_K110003:
 http://baderlab.bme.jhu.edu:8888/moodle18/mod/data/view.php?d=4&rid=1462
 http://baderlab.bme.jhu.edu:8888/moodle18/mod/data/view.php?d=4&rid=1463
 BBa_K110005, BBa_K110006:
 http://baderlab.bme.jhu.edu:8888/moodle18/mod/data/view.php?d=4&rid=1470
 Cloning of PCR product successful: Y/N  Yes 
 Sequencing of cloned PCR product successful: Y/N  No
 Joining of validated part to adjacent part(s) status:  Not Done
 Problems to be solved: Not really a problem, but need do a mini-Prep and sequence
 Current status of this part: All cloned and inoculated into 1.5 mL LB for mini-prep.

GROUP 5: MATa Specific Promoters II

 Date: 7/11/08
 status report by Rick Carrick
 Part no.: BBa_K110015,
 Part Description: MFA1
 Part Location (in build a genome lab):
 PCR successful?; Y (on moodle somewhere)
 Cloning of PCR product successful: Y
 Sequencing of cloned PCR product successful:N
 Joining of validated part to adjacent part(s) status: Not done
 Problems to be solved: None so far
 Current status of this part: This parts must be restriction enzyme digested and sequenced 
 next.
 Date: 7/11/08
 status report by Rick Carrick
 Part no.: BBa_K110009
 Part Description: Ste2
 Part Location (in build a genome lab):
 PCR successful?; Y (on moodle somewhere)
 Cloning of PCR product successful: Y
 Sequencing of cloned PCR product successful:N
 Joining of validated part to adjacent part(s) status: Not done
 Problems to be solved: None so far
 Current status of this part: This part must be restriction enzyme digested and sequenced 
next

GROUP 6: Vectors

 Status report by ____
 Vector transformed into bacteria strain DB3.1 Y/N
 Permanent culture made in the Boeke lab for future reference Y/N
 Selectable marker for this vector
 Medium made and tested Y/N  (link)
 DNA preps made Y/N
 DNA preps tested by RE digest  - (link)
 DNA preps tested by transformation into DB3.1 and DH5alpha (or JM109) – put data as 
   Table on moodle.  
 Sample format/data follows:
Amount transformed cfu/micGm in DB3.1 cfu/micGm in JM109 0.1 ng 5 * 10e7 <2 * 10e2
Preparative digests ready for use are located – where?

GROUP 7: Microscopy/Yeast

 Milestones 
 Have gfp yeast been visualized by microscopy?
 Have gfp yeast been photographed? Link to moodle – picture.  
 [If pics are nice, put on our web page]
 Can green(/other colors) colonies be photographed? 
 Has STE3-gfp sex detector been transformed into MATa, MATalpha, and MATa/alpha?  Y/N
 Have permanent cultures been banked in the Boeke lab? Y/N
 Have STE3-gfp sex detector cells/colonies been photographed Y/N  - link to Moodle