LOCUS       40924_31980        8035 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Shuttle vector pMQ122, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8035)
  AUTHORS   Shanks RMQ., MacEachran DP, O'Toole GA.
  TITLE     In vivo recombination vectors for modification and expression of 
            Gram-negative and Gram-positive bacterial genes
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8035)
  AUTHORS   Shanks RMQ., MacEachran DP, O'Toole GA.
  TITLE     Direct Submission
  JOURNAL   Submitted (12-FEB-2008) Ophthalmololgy, University of Pittsburgh, 
            203 Lothrop st, Pittsburgh, PA 15213, USA
REFERENCE   3  (bases 1 to 8035)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8035)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (12-FEB-2008) Ophthalmololgy, University of Pittsburgh, 203 Lothrop 
            st, Pittsburgh, PA 15213, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..8035
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..801
                     /codon_start=1
                     /label=URA3
                     /note="orotidine-5'-phosphate decarboxylase, required for
                     uracil biosynthesis"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     oriT            complement(893..1001)
                     /direction=LEFT
                     /label=oriT
                     /note="incP origin of transfer"
     protein_bind    complement(1198..1231)
                     /label=loxP
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     rep_origin      complement(1282..1633)
                     /direction=LEFT
                     /label=pRO1600 oriV
                     /note="broad-host-range origin of replication from
                     Pseudomonas aeruginosa plasmid pRO1600; requires the 
                     pRO1600 Rep protein for replication (West et al., 1994)"
     CDS             1647..2477
                     /codon_start=1
                     /label=pRO1600 Rep
                     /note="replication protein for the broad-host-range plasmid
                     pRO1600 from Pseudomonas aeruginosa"
                     /translation="VASPPMVYKSNALVEAAYRLSVQEQRIVLACISQVKRSEPVTDEV
                     MYSVTAEDIATMAGVPIESSYNQLKEAALRLKRREVRLTQEPNGKGKRPSVMITGWVQT
                     IIYREGEGRVELRFTKDMLPYLTELTKQFTKYALADVAKMDSTHAIRLYELLMQWDSIG
                     QREIEIDQLRKWFQLEGRYPSIKDFKLRVLDPAVTQINEHSPLQVEWAQRKTGRKVTHL
                     LFSFGPKKPAKAVGKAPAKRKAGKISDAEIAKQARPGETWEAARARLTQMPLDLA"
     terminator      complement(2679..2765)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     CDS             complement(2995..3708)
                     /codon_start=1
                     /label=yeGFP
                     /note="yeast-enhanced green fluorescent protein"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     protein_bind    complement(3807..3823)
                     /label=lac operator
                     /note="The lac repressor binds to the lac operator to
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(3831..3859)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and
                     lac UV5 promoters"
     promoter        4089..4166
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             4290..5105
                     /codon_start=1
                     /gene="lacIQ"
                     /product="truncated LacIQ"
                     /label=lacIQ
                     /protein_id="ACA49554.1"
                     /translation="MAELNYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSR
                     ADQLGASVVVSMVERSGVEACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPA
                     LFLDVSDQTPINSIIFSHEDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKY
                     LTRNQIQPIAEREGDWSAMSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLR
                     VGADISVVGYDDTEDSSCYIPPLTTIKQDFRLLGQTTWTACCNSLRARR"
     gene            4290..5105
                     /gene="lacIQ"
                     /label=lacIQ
     rep_origin      complement(5286..5874)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(6048..6905)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(6906..7010)
                     /label=AmpR promoter
     misc_feature    7047..7550
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an
                     autonomously replicating sequence"
     promoter        7815..8035
                     /label=URA3 promoter
ORIGIN
        1 atgtcgaaag ctacatataa ggaacgtgct gctactcatc ctagtcctgt tgctgccaag
       61 ctatttaata tcatgcacga aaagcaaaca aacttgtgtg cttcattgga tgttcgtacc
      121 accaaggaat tactggagtt agttgaagca ttaggtccca aaatttgttt actaaaaaca
      181 catgtggata tcttgactga tttttccatg gagggcacag ttaagccgct aaaggcatta
      241 tccgccaagt acaatttttt actcttcgaa gacagaaaat ttgctgacat tggtaataca
      301 gtcaaattgc agtactctgc gggtgtatac agaatagcag aatgggcaga cattacgaat
      361 gcacacggtg tggtgggccc aggtattgtt agcggtttga agcaggcggc agaagaagta
      421 acaaaggaac ctagaggcct tttgatgtta gcagaattgt catgcaaggg ctccctatct
      481 actggagaat atactaaggg tactgttgac attgcgaaga gcgacaaaga ttttgttatc
      541 ggctttattg ctcaaagaga catgggtgga agagatgaag gttacgattg gttgattatg
      601 acacccggtg tgggtttaga tgacaaggga gacgcattgg gtcaacagta tagaaccgtg
      661 gatgatgtgg tctctacagg atctgacatt attattgttg gaagaggact atttgcaaag
      721 ggaagggatg ctaaggtaga gggtgaacgt tacagaaaag caggctggga agcatatttg
      781 agaagatgcg gccagcaaaa ctaaaaaact gtattataag taaatgcatg tatactaaac
      841 tcacaaatta gagcttcaat ttaattatat cagttattca cgtggagctt atcggccagc
      901 ctcgcagagc aggattcccg ttgagcaccg ccaggtgcga ataagggaca gtgaagaagg
      961 aacacccgct cgcgggtggg cctacttcac ctatcctgcc cggctgacgc cgttggatac
     1021 accaaggaaa gtctacacga accctttggc aaaatcctgt atatcgtgcg aaaaaggatg
     1081 gatataccga aaaaatcgct ataatgaccc cgaagcaggg ttatgcagcg gaaagtatac
     1141 cttaaccgcc ggctgacccg gcggggacga ggcaagctaa acagatctct agacctaata
     1201 acttcgtata gcatacatta tacgaagtta tattaagggt tgtcgagccg ctggtgccgc
     1261 tggttggacg ccaagggtga atccgcctcg ataccctgat tactcgcttc ctgcgccctc
     1321 tcaggcggcg ataggggact ggtaaaacgg ggattgccca gacgcctccc ccgccccttc
     1381 aggggcacaa atgcggcccc aacggggcca cgtagtggtg cgttttttgc gtttccaccc
     1441 ttttcttcct tttccctttt aaacctttta ggacgtctac aggccacgta atccgtggcc
     1501 tgtagagttt aaaaagggac ggatttgttg ccattaaggg acggatttgt tgttaagaag
     1561 ggacggattt gttgttgtaa agggacggat ttgttgtatt gtgggacgca gatacagtgt
     1621 ccccttatac acaaggaatg tcgaacgtgg cctcaccccc aatggtttac aaaagcaatg
     1681 ccctggtcga ggccgcgtat cgcctcagtg ttcaggaaca gcggatcgtt ctggcctgta
     1741 ttagccaggt gaagaggagc gagcctgtca ccgatgaagt gatgtattca gtgacggcgg
     1801 aggacatagc gacgatggcg ggtgtcccta tcgaatcttc ctacaaccag ctcaaagaag
     1861 cggccctgcg cctgaaacgg cgggaagtcc ggttaaccca agagcccaat ggcaagggga
     1921 aaagaccgag tgtgatgatt accggctggg tgcaaacaat catctaccgg gagggtgagg
     1981 gccgtgtaga actcaggttc accaaagaca tgctgccgta cctgacggaa ctcaccaaac
     2041 agttcaccaa atacgccttg gctgacgtgg ccaagatgga cagcacccac gcgatcaggc
     2101 tttacgagct gctcatgcaa tgggacagca tcggccagcg cgaaatagaa attgaccagc
     2161 tgcgaaagtg gtttcaactg gaaggccggt atccctcgat caaggacttc aagttgcgag
     2221 tgcttgatcc agccgtgacg cagatcaacg agcacagccc gctacaggtg gagtgggcgc
     2281 agcgaaagac cgggcgcaag gtcacacatc tgttgttcag ttttggaccg aagaagcccg
     2341 ccaaggcggt gggtaaggcc ccagcgaagc gcaaggccgg gaagatttca gatgctgaga
     2401 tcgcgaaaca ggctcgccct ggtgagacat gggaagcggc ccgcgctcga ctaacccaga
     2461 tgccgctgga tctggcctag aggccgtggc caccacggcc cggcctgcct ttcaggctgc
     2521 gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag
     2581 ggggatgtgc tgcaaggatg cctggcagtt tatggcgggc gtcctgcccg ccaccctccg
     2641 ggccgttgct tcgcaacgtt caaatccgct cccggcggat ttgtcctact caggagagcg
     2701 ttcaccgaca aacaacagat aaaacgaaag gcccagtctt tcgactgagc ctttcgtttt
     2761 atttgatgcc tggcagttcc ctactctcgc atggggagac cccacactac catcggcgct
     2821 acggcgtttc acttctgagt tcggcatggg gtcaggtggg accaccgcgc tactgccgcc
     2881 aggcaaattc tgttttatca gaccgcttct gcgttctgat ttaatctgta tcaggctgaa
     2941 aatcttctct catccgccaa aacagccaag cttgcatgcc tgcagactag tctatttgta
     3001 tagttcatcc atgccatgtg taatcccagc agctgttaca aactcaagaa ggaccatgtg
     3061 gtctctcttt tcgttgggat ctttcgaaag ggcagattgt gtggacaggt aatggttgtc
     3121 tggtaaaagg acagggccat cgccaattgg agtattttgt tgataatggt ctgctagttg
     3181 aacgcttcca tcttcaatgt tgtgtctaat tttgaagtta actttgattc cattcttttg
     3241 tttgtctgcc atgatgtata cattgtgtga gttatagttg tattccaatt tgtgtccaag
     3301 aatgtttcca tcttctttaa aatcaatacc ttttaactcg attctattaa caagggtatc
     3361 accttcaaac ttgacttcag cacgtgtctt gtagttcccg tcatctttga aaaatatagt
     3421 tctttcctgt acataacctt cgggcatggc actcttgaaa aagtcatgct gtttcatatg
     3481 atctgggtat ctcgcaaagc attgaacacc ataaccgaaa gtagtgacaa gtgttggcca
     3541 tggaacaggt agttttccag tagtgcaaat aaatttaagg gtaagttttc cgtatgttgc
     3601 atcaccttca ccctctccac tgacagaaaa tttgtgccca ttaacatcac catctaattc
     3661 aacaagaatt gggacaactc cagtgaaaag ttcttctcct ttactcatat gtatatctcc
     3721 ttccccgggt accgagctcg aattcgctag cccaaaaaaa cgggtatgga gaatctagag
     3781 gatccaattc tgtttcctgt gtgaaattgt tatccgctca caattccaca cattatacga
     3841 gccgatgatt aattgtcaac agctcatttc agaatatttg ccagaaccgt tatgatgtcg
     3901 gcgcaaaaaa cattatccag aacgggagtg cgccttgagc gacacgaatt atgcagtgat
     3961 ttacgacctg cacagccata ccacagcttc cgatggctgc ctgacgccag aagcattggt
     4021 gcaccgtgca gtcgataagc ccggatcagc ttgcaattcg cgcgcgaagg cgaagcggca
     4081 tttacgttga caccatcgaa tggtgcaaaa cctttcgcgg tatggcatga tagcgcccgg
     4141 aagagagtca attcagggtg gtgaatgtga aaccagtaac gttatacgat gtcgcagagt
     4201 atgccggtgt ctcttatcag accgtttccc gcgtggtgaa ccaggccagc cacgtttctg
     4261 cgaaaacgcg ggaaaaagtg gaagcggcga tggcggagct gaattacatt cccaaccgcg
     4321 tggcacaaca actggcgggc aaacagtcgt tgctgattgg cgttgccacc tccagtctgg
     4381 ccctgcacgc gccgtcgcaa attgtcgcgg cgattaaatc tcgcgccgat caactgggtg
     4441 ccagcgtggt ggtgtcgatg gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc
     4501 acaatcttct cgcgcaacgc gtcagtgggc tgatcattaa ctatccgctg gatgaccagg
     4561 atgccattgc tgtggaagct gcctgcacta atgttccggc gttatttctt gatgtctctg
     4621 accagacacc catcaacagt attattttct cccatgaaga cggtacgcga ctgggcgtgg
     4681 agcatctggt cgcattgggt caccagcaaa tcgcgctgtt agcgggccca ttaagttctg
     4741 tctcggcgcg tctgcgtctg gctggctggc ataaatatct cactcgcaat caaattcagc
     4801 cgatagcgga acgggaaggc gactggagtg ccatgtccgg ttttcaacaa accatgcaaa
     4861 tgctgaatga gggcatcgtt cccactgcga tgctggttgc caacgatcag atggcgctgg
     4921 gcgcaatgcg cgccattacc gagtccgggc tgcgcgttgg tgcggatatc tcggtagtgg
     4981 gatacgacga taccgaagac agctcatgtt atatcccgcc gttaaccacc atcaaacagg
     5041 attttcgcct gctggggcaa accacgtgga ccgcttgctg caactctctc agggccaggc
     5101 ggtgaagggc aatcagctgt tgcccgtctc actggctcgg tcgttcggct gcggcgagcg
     5161 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     5221 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     5281 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     5341 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     5401 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     5461 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     5521 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     5581 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     5641 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     5701 tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca
     5761 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     5821 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     5881 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     5941 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     6001 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     6061 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     6121 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     6181 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     6241 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     6301 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     6361 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     6421 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     6481 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     6541 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     6601 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     6661 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     6721 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     6781 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     6841 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     6901 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     6961 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     7021 cgaaaagtgc cacctgggtc cttttcatca cgtgctataa aaataattat aatttaaatt
     7081 ttttaatata aatatataaa ttaaaaatag aaagtaaaaa aagaaattaa agaaaaaata
     7141 gtttttgttt tccgaagatg taaaagactc tagggggatc gccaacaaat actacctttt
     7201 atcttgctct tcctgctctc aggtattaat gccgaattgt ttcatcttgt ctgtgtagaa
     7261 gaccacacac gaaaatcctg tgattttaca ttttacttat cgttaatcga atgtatatct
     7321 atttaatctg cttttcttgt ctaataaata tatatgtaaa gtacgctttt tgttgaaatt
     7381 ttttaaacct ttgtttattt ttttttcttc attccgtaac tcttctacct tctttattta
     7441 ctttctaaaa tccaaataca aaacataaaa ataaataaac acagagtaaa ttcccaaatt
     7501 attccatcat taaaagatac gaggcgcgtg taagttacag gcaagcgatc cgtcctaaga
     7561 aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc cctttcgtct
     7621 cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg agacggtcac
     7681 agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt
     7741 tggcgggtgt cggggctggc ttaactatgc ggcatcagag cagattgtac tgagagtgca
     7801 ccataccaca gcttttcaat tcaattcatc attttttttt tattcttttt tttgatttcg
     7861 gtttctttga aatttttttg attcggtaat ctccgaacag aaggaagaac gaaggaagga
     7921 gcacagactt agattggtat atatacgcat atgtagtgtt gaagaaacat gaaattgccc
     7981 agtattctta acccaactgc acagaacaaa aacctgcagg aaacgaagat aaatc
//
