Team:Johns Hopkins/Notebook/GROUP 4: Long Two-way Stops & Mat(alpha) specific promotors

GROUP 4: Long Two-way Stops & Mat(alpha) specific promoters
Summary: As of September 23rd, 2008 We had successfully cloned BBa_K110005 and BBa_K110006 (alpha Promoters) and BBa_K110003 (Long Terminator) into pGEM-T and Kanamycin biobrick vectors. They await assembly and ligation.

Date: September 17th, 2008 Minipreps were made of the kanamycin biobrick vector clones, and digested with E/P to  check for insert of correct size. See gel: /pic// Low-copy origin makes for poor gel visualization. Suggest some other diagnostic test. Date: September 12th, 2008 Clones were digested with E/P, and ligated O/N into Kanamycin biobrick vector (pSB4K5). Colonies were picked and grown.

Date: August 26th, 2008 Author: Jonathan Sequences were returned, and prefix/suffix/identities were confirmed. Clones that passed are: Long Terminator BBa_K110003: Clone A1-3.2 Alpha Promoter BBa_K110005: Clone 5.1 Alpha Promoter BBa_K110006: Clones 6.1, 6.3

Date: August 21st, 2008 Author: Jonathan Clones were grown up O/N and re-miniprepped. Digestion with E/P yielded some possible candidates. Here is the gel pic: 08/21/08 E/P Restriction Digest for BBs 3, 5, 6 (pGEM-T) Clones submitted for sequencing.

Date: August 1st, 2008 Author: Jonathan Sequences were submitted to the facility (3/building block) and a few returned perfect clones, but due to miscommunication, exactly which clones that the trace files correspond to are under some confusion. Will attempt to confirm clone identities.

Date: July 29, 2008 Status report by: Jaime Part no.: BBa_K110001, BBa_K110003, BBa_K110005, BBa_K110006 Part Description: Long Two-way Stops & Mat(alpha) specific promotors Restriction Enzyme Digest of Mini-Preps

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 iGEM PCR experimentation on BBa_K110001 and BBa_K110003 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 iGEM PCR experimentation 2 on BBa_K110001 and BBa_K110003 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 iGEM BBa_K110005 and BBa_K110006 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 iGEM BBa_K110005 and BBa_K110006 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