LOCUS       40924_4644        7938 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pAP309, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7938)
  AUTHORS   Park AS, Rubin EJ.
  TITLE     Clp-mediated Regulation of WhiB1 is Required for Cell Division in 
            Mycobacteria
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7938)
  AUTHORS   Park AS, Rubin EJ.
  TITLE     Direct Submission
  JOURNAL   Submitted (11-JUL-2017) Department of Immunology and Infectious 
            Diseases, Harvard School of Public Health, 665 Huntington Ave, 
            Boston, MA 02115, USA
REFERENCE   3  (bases 1 to 7938)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 7938)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (11-JUL-2017) Department of Immunology and Infectious Diseases, 
            Harvard School of Public Health, 665 Huntington Ave, Boston, MA 
            02115, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..7938
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      20..47
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     regulatory      72..377
                     /regulatory_class="promoter"
     protein_bind    247..264
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     misc_feature    406..657
                     /note="Region: WhiB family transcriptional regulator with 
                     mutation (R73E)"
     CDS             679..1392
                     /label=EGFP
                     /note="enhanced GFP"
     CDS             1393..1410
                     /label=6xHis
                     /note="6xHis affinity tag"
     CDS             1429..1449
                     /label=TEV site
                     /note="tobacco etch virus (TEV) protease recognition and 
                     cleavage site"
     CDS             1456..1473
                     /label=6xHis
                     /note="6xHis affinity tag"
     CDS             1480..1509
                     /label=Myc
                     /note="Myc (human c-Myc proto-oncogene) epitope tag"
     regulatory      1534..1576
                     /regulatory_class="terminator"
     primer_bind     complement(1579..1595)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     terminator      4420..4447
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     CDS             4631..5251
                     /label=TetR
                     /note="tetracycline repressor TetR"
     primer_bind     5867..5883
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    complement(5918..6052)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(6119..6490)
                     /label=BleoR
                     /note="antibiotic-binding protein"
     primer_bind     complement(6657..6673)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(6681..6697)
                     /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(6705..6735)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(6750..6771)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      7109..7697
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 actagttgat tagctaagca gaaggccatc ctgacggatg gcctttttgc gtttaatact
       61 gtttaaactc tagaaatatt ggatcgtcgg caccgtcacg gccgtgggag gcggcacgat
      121 ccgcgacgtg atgatcggcc gcatccccac ggtgctgcgc agtgagctct acgccatccc
      181 ggcgttgatc tgtgcgttcg cacgcacagg cccggtgtga gaagggtctc tgacgagcgg
      241 gagaactccc tatcagtgat agagtttgtc ctccctatca gtgatagata ggctctggga
      301 gtacccgtgt gtacgaccag cacggcatac atcatttcga cgccgagaga ttcgccgccc
      361 gaaatgagca cgatccgcat gcttaattaa gaaggagata tacatatgga ttggcgccac
      421 aaggcggtct gtcgcgacga ggatccggag ctgttcttcc cggtgggaaa cagtgggccg
      481 gcccttgccc agatcgctga cgcgaagctg gtgtgcaacc ggtgcccggt gaccaccgaa
      541 tgcctgagct gggctttgga gtcgggccag gacgccggcg tgtggggcgg catgagcgag
      601 gacgagcgcc gtgcactcaa ggagcgcaac gcgcgcacca aggcgcgcac cggagtcggc
      661 ggctcggaat tcggtaccgt gtcgaagggc gaggagctgt tcaccggcgt cgtgcccatc
      721 ctggtcgagc tggacggcga cgtgaacggc cacaagttca gcgtctcggg cgagggcgag
      781 ggcgacgcga cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccg
      841 gtcccctggc cgaccctggt gaccaccctg acctacggcg tgcagtgttt ctcgcggtac
      901 cccgaccaca tgaagcagca cgacttcttc aagagcgcga tgccggaggg ctacgtccag
      961 gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgggccga ggtcaagttc
     1021 gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaagacggc
     1081 aacatcctgg gccacaagct ggagtacaac tacaactcgc acaacgtcta catcatggcg
     1141 gacaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc
     1201 agcgtccagc tggccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtcctg
     1261 ctgccggaca accactacct gagcacccag tcggcgctga gcaaggaccc gaacgagaag
     1321 cgggaccaca tggtcctgct ggagttcgtg accgccgcgg gcatcaccct gggcatggac
     1381 gagctgtaca agcaccacca ccaccaccat taggatatcc atttaaatga gaacctgtac
     1441 ttccagggca ctagtcacca ccaccaccac catatggccg agcagaagct gatctcggag
     1501 gaggacctgt gatctagaat acgcatcaag ctttagctaa ttaattgggg accctagagg
     1561 tccccttttt tattttatcg ataccgtcga cctcgagggg gggcccgccg gggatcatcg
     1621 agccgagaac gttatcgaag ttggtcatgt gtaatcccct cgtttgaact ttggattaag
     1681 cgtagataca cccttggaca agccagttgg attcggagac aagcaaattc agccttaaaa
     1741 agggcgaggc ctgcggtggt ggaacaccgc agggcctcta accgctcgac gcgctgcacc
     1801 aaccagcccg cgaacggctg gcagccagcg taaggcgcgg ctcatcgggc ggcgttcgcc
     1861 acgatgtcct gcacttcgag ccaagcctcg aacacctgct ggtgtgcacg actcacccgg
     1921 ttgttgacac cgcgcgcggc cgtgcgggct cggtggggcg gctgtgtcgc ccttgccagc
     1981 gtgagtagcg cgtacctcac ctcgcccaac aggtcgcaca cagccgattc gtacgccata
     2041 aagccaggtg agcccaccag ctccgtaagt tcgggcgctg tgtggctcgt acccgcgcat
     2101 tcaggcggca gggggtctaa cgggtctaag gcggcgtgta cggccgccac agcggctctc
     2161 agcggcccgg aaacgtcctc gaaacgacgc atgtgttcct cctggttggt acaggtggtt
     2221 gggggtgctc ggctgtcgct ggtgttccac caccagggct cgacgggaga gcgggggagt
     2281 gtgcagttgt ggggtggccc ctcagcgaaa tatctgactt ggagctcgtg tcggaccata
     2341 caccggtgat taatcgtggt ctactaccaa gcgtgagcca cgtcgccgac gaatttgagc
     2401 agctctggct gccgtactgg ccgctggcaa gcgacgatct gctcgagggg atctaccgcc
     2461 aaagccgcgc gtcggcccta ggccgccggt acatcgaggc gaacccaaca gcgctggcaa
     2521 acctgctggt cgtggacgta gaccatccag acgcagcgct ccgagcgctc agcgcccggg
     2581 ggtcccatcc gctgcccaac gcgatcgtgg gcaatcgcgc caacggccac gcacacgcag
     2641 tgtgggcact caacgcccct gttccacgca ccgaatacgc gcggcgtaag ccgctcgcat
     2701 acatggcggc gtgcgccgaa ggccttcggc gcgccgtcga cggcgaccgc agttactcag
     2761 gcctcatgac caaaaacccc ggccacatcg cctgggaaac ggaatggctc cactcagatc
     2821 tctacacact cagccacatc gaggccgagc tcggcgcgaa catgccaccg ccgcgctggc
     2881 gtcagcagac cacgtacaaa gcggctccga cgccgctagg gcggaattgc gcactgttcg
     2941 attccgtcag gttgtgggcc tatcgtcccg ccctcatgcg gatctacctg ccgacccgga
     3001 acgtggacgg actcggccgc gcgatctatg ccgagtgcca cgcgcgaaac gccgaattcc
     3061 cgtgcaacga cgtgtgtccc ggaccgctac cggacagcga ggtccgcgcc atcgccaaca
     3121 gcatttggcg ttggatcaca accaagtcgc gcatttgggc ggacgggatc gtggtctacg
     3181 aggccacact cagtgcgcgc cagtcggcca tctcgcggaa gggcgcagca gcgcgcacgg
     3241 cggcgagcac agttgcgcgg cgcgcaaagt ccgcgtcagc catggaggca ttgctatgag
     3301 cgacggctac agcgacggct acagcgacgg ctacaaccgg cagccgactg tccgcaaaaa
     3361 gcggcgcgtg accgccgccg aaggcgctcg aatcaccgga ctatccgaac gccacgtcgt
     3421 ccggctcgtg gcgcaggaac gcagcgagtg gctcgccgag caggctgcac gccgcgaacg
     3481 catccgcgcc tatcacgacg acgagggcca ctcttggccg caaacggcca aacatttcgg
     3541 gctgcatctg gacaccgtta agcgactcgg ctatcgggcg aggaaagagc gtgcggcaga
     3601 acaggaagcg gctcaaaagg cccacaacga agccgacaat ccaccgctgt tctaacgcaa
     3661 ttggggagcg ggtgtcgcgg gggttccgtg gggggttccg ttgcaacggg tcggacaggt
     3721 aaaagtcctg gtagacgcta gttttctggt ttgggccatg cctgtctcgt tgcgtgtttc
     3781 gttgcgcccg ttttgaatac cagccagacg agacggggtt ctacgaatct tggtcgatac
     3841 caagccattt ccgctgaata tcggggagct caccgccaga atcggtggtt gtggtgatgt
     3901 acgtggcgaa ctccgttgta gtgcctgtgg tggcatccgt ggccactctc gttgcacggt
     3961 tcgttgtgcc gttacaggcc ccgttgacag ctcaccgaac gtagttaaaa catgctggtc
     4021 aaactaggtt taccaacgat acgagtcagc tcatctaggg ccagttctag gcgttgttcg
     4081 ttgcgcggtt cgttgcgcat gtttcgtgtg gttgctagat ggctccgcaa ccacacgctt
     4141 cgaggttgag tgcttccagc acgggcgcga tccagaagaa cttcgtcgtg cgactgtcct
     4201 cgttgatcct tgccgagctg ggatggaagc tcggccgacc accctggagg agatgatcga
     4261 ggatgccagg gcctttcacg cccgccgctg ctgagcgtcc gccgccgggc ccgcaccgcc
     4321 gtcggccggc ccgctccggg ctcgcagcag cgggcttcgg cgcgggcccg gggctcccga
     4381 gcgcgggcgg ggctccgggc ggccgctgat tagctaagca gaaggccatc ctgacggatg
     4441 gcctttttgc gtttaatact gcatgcactc tagaaatatt ggatcgtcgc accgggttaa
     4501 gcctggagtg cggtggtgcc tggtcggcat tttcgcagtc gagggctctc gtgtagcctg
     4561 ggcgagttgc cgacgcaggc gaccctctgc cacggatccg catggtacca tcaggaggaa
     4621 tcacttcgca atgtctagat tagataaaag taaagtgatt aacagcgcat tagagctgct
     4681 taatgaggtc ggaatcgaag gtttaacaac ccgtaaactc gcccagaagc ttggtgtaga
     4741 gcagcctaca ttgtattggc atgtaaaaaa taagcgggcc ctgctcgacg ccttagccat
     4801 tgagatgtta gataggcacc atactcactt ttgcccttta gaaggggaaa gctggcaaga
     4861 ttttttacgt aataacgcta aaagttttag atgtgcttta ctaagtcatc gcgatggagc
     4921 aaaagtacat ttaggtacac ggcctacaga aaaacagtat gaaactctcg aaaatcaatt
     4981 agccttttta tgccaacaag gtttttcact agagaacgcg ttatatgcac tcagcgctgt
     5041 ggggcatttt accttaggtt gcgtattgga agatcaagag catcaagtcg ctaaagaaga
     5101 aagggaaaca cctactactg atagtatgcc gccattatta cgacaagcta tcgaattatt
     5161 tgatcaccaa ggtgcagagc cagccttctt attcggcctt gaattgatca tatgcggatt
     5221 agaaaaacaa cttaaatgtg aaagtgggtc ttaatgacca tgggctagcg gctttagcta
     5281 attaattggg gaccctagag gtcccctttt ttattttgcg gccgccgggg gccgggggcg
     5341 gcgccgggcg gcccggggcg tcaggcgccg ggggcggtgt ccggcggccc ccagaggaac
     5401 tgcgccagtt cctccggatc ggtgaagccg gagagatcca gcggggtctc ctcgaacacc
     5461 tcgaagtcgt gcaggaaggt gaaggcgagc agttcgcggg cgaagtcctc ggtccgcttc
     5521 cactgcgccc cgtcgagcag cgcggccagg atctcgcggt cgccccggaa ggcgttgaga
     5581 tgcagttgca ccaggctgta gcgggagtct cccgcataga cgtcggtgaa gtcgacgatc
     5641 ccggtgacct cggtcgcggc caggtccacg aagatgttgg tcccgtgcag gtcgccgtgg
     5701 acgaaccggg gttcgcggcc ggccagcagc gtgtccacgt ccggcagcca gtcctccagg
     5761 cggtccagca gccggggcga gaggtagccc cacccgcggg cgaaaggggg atgtgctgca
     5821 aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa aacgacggcc
     5881 agtgccaagc ttgcatgcct gcaggtcgac tctagaggat ccagacatga taagatacat
     5941 tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta tttgtgaaat
     6001 ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag tttcgaggtc
     6061 gacccccctg ggacccgtgg gccgccgtcg gaccggcggt gttggtcggc gtcggtcagt
     6121 cctgctcctc ggccacgaag tgcacgcagt tgccggccgg gtcgcgcagg gcgaactccc
     6181 gcccccacgg ctgctcgccg atctcggtca tggccggccc ggaggcgtcc cggaagttcg
     6241 tggacacgac ctccgaccac tcggcgtaca gctcgtccag gccgcgcacc cacacccagg
     6301 ccagggtgtt gtccggcacc acctggtcct ggaccgcgct gatgaacagg gtcacgtcgt
     6361 cccggaccac accggcgaag tcgtcctcca cgaagtcccg ggagaacccg agccggtcgg
     6421 tccagaactc gaccgctccg gcgacgtcgc gcgcggtgag caccggaacg gcactggtca
     6481 acttggccat ggtgaatcct cctgaatatg tagagcttct ttcctgcact atgatgagtg
     6541 caccccgctc aagcttgcct cgacggcggt gtgctgtcaa ttcagagctg agcctgatgc
     6601 actcaactta ctgagcatgc taacgctggt cgtgcgggtc ggaattcgta atcatggtca
     6661 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     6721 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     6781 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc cgcggtcagg cgttgacgat
     6841 gacgggctgg tcggccacgt ccggggagtc aggcaactat ggatgaacga aatagacaga
     6901 tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat
     6961 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     7021 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     7081 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     7141 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     7201 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc
     7261 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     7321 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     7381 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     7441 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     7501 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     7561 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     7621 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     7681 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     7741 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
     7801 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag
     7861 tgagcgagga agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga
     7921 ttcattaatg cagctaga
//
