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Team:Johns Hopkins/Notebook
From 2008.igem.org
Contents |
Groups
iGEM Groups 1.0
iGEM Groups 2.01
iGEM Groups 2.02
Important reminders and notes
[Can make general comments here, so they don't get lost in peoples e-mail boxes] July 11: Primers for group 1 were delivered yesterday. July 11: Lab meeting at 7:30PM in the lab to go over miniprep protocol. July 15: Lab meeting at 6:30PM with Jessica. Have status reports ready. Bring labtop if you can. July 17: Restriction Digest/Sequencing Preparation (with James) 6:00PM. July 21: 6:00PM or 6:30PM Lab meeting with Jessica. Have status reports ready.
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
Status Report 2.2 - 07/17/08
Please BOLD the most recent step that you have completed. Do this by placing the tag < b > in front of and </ b > at the end of what you would like to be placed in bold (with no space between letter and carrot symbol (<,>).
GROUP 1: Fluorescent Proteins
Summary for Fluorescent Proteins Group
Date: July 22, 2008 Status report by: Tejas Part no.: BBa_K110017 -> BBa_K110023 Part Description: yESapphire , mCherry, venusYFP, and Citrine Work on yESapphire was gratiously done by James. Primers were designed. Restriction site ends have been added through PCR. The product was cloned into JM109. Colonies were picked and an inoculation was grown. Miniprep was performed and subsequent DNA was CS PCR'd and run on a gel to verify contents. BioBrick is currently being sequenced. Work on mCherry and venusYFP (BBa_K110018 -> BBa_K110021) is currently being done. Primers were designed. Restriction sites have been added through PCR. The products were cloned into JM109 and plated. 3 colonies per 4 BioBricks (total 12) were picked, grown out, miniprepped, and digested for verification on a gel. According to the results, (Venus YFP and mCherry Miniprep check via digest), only one of the mCherry's (BBa_K110019) is the correct product. This product will be retransformed to yield more for sequencing. Colonies for the other three BioBricks were selected by Ingrid on 7.22.2008 and are being grown out. Process will continue as per protocol. Work on Citrine has not yet begun. Primers have been designed and ordered, but template DNA must be grown out. Template DNA to be grown and extracted by James should we later decide that Citrine will be needed. *Note that mCherry and Citrine both have restriction sites within the coding region, and are therefore not optimal. Advice/help on this issue would be appreciated.
GROUP 2: MATa Specific-promoters
Summary for MATa Specific Promoters Group
Date: _________ __, 2008 Status report by: _____ Part no.: BBa_K1100XX -> BBa_K1100YY Part Description: Summary here.
GROUP 3: Short Two-Way Stops
Summary for Short Two-Way Stops Group
Date: _________ __, 2008 Status report by: _____ Part no.: BBa_K1100XX -> BBa_K1100YY Part Description: Summary here.
GROUP 4: Long Two-Way Stops & MAT(alpha) Specific Promotors
Date: 7/10/08 Status report by: Jaime Liu Part no.: BBa_K110001 Part Description: BBa_K110001 - Between-bud 27-W FRS2-C + 200bp into each gene LtR Part Location (in build a genome lab): In 4C fridge #2 PCR successful?; Y/N (link such as this)- Yes BBa_K110001: http://baderlab.bme.jhu.edu:8888/moodle18/mod/data/view.php?d=4&rid=1462 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 100uL in 96 well plate for sequencing- put in incubator on 7/15
Date: 7/10/08 Status report by: Jaime Liu Part no.: BBa_K110003 Part Description: BBa_K110003 - Between-SWP82-W and EMP47-C +200 into each gene LtR Part Location (in build a genome lab): In 4C fridge #2 PCR successful?; Y/N (link such as this)- Yes BBa_K110003: http://baderlab.bme.jhu.edu:8888/moodle18/mod/data/view.php?d=4&rid=1463 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 100uL in 96 well plate for sequencing- put in incubator on 7/15
Date: 7/10/08 Status report by: Jaime Liu Part no.: BBa_K110005 Part Description: BBa_K110005 - MFalpha2 LtR Part Location (in build a genome lab): In 4C fridge #2 PCR successful?; Y/N (link such as this)- Yes BBa_K110005: 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 100uL in 96 well plate for sequencing- put in incubator on 7/15
Date: 7/10/08 Status report by: Jaime Liu Part no.: BBa_K110006 Part Description: 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_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 100uL in 96 well plate for sequencing- put in incubator on 7/15
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
