"
Team:Johns Hopkins/Notebook
From 2008.igem.org
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
