LOCUS       62056_10195        7372 bp DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7372)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..7372
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(140..157)
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     CDS             complement(2210..2242)
                     /codon_start=1
                     /label=T7 tag (gene 10 leader)
                     /note="leader peptide from bacteriophage T7 gene 10"
                     /translation="MASMTGGQQMG"
     CDS             complement(2246..2263)
                     /codon_start=1
                     /label=thrombin site
                     /note="thrombin recognition and cleavage site"
                     /translation="LVPRGS"
     CDS             complement(2273..2290)
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     RBS             complement(2309..2331)
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    complement(2346..2370)
                     /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(2371..2389)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        2698..2775
                     /label=lacI promoter
     CDS             2776..3855
                     /codon_start=1
                     /label=lacI
                     /note="lac repressor"
                     /translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     protein_bind    3871..3892
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             4667..4855
                     /codon_start=1
                     /label=rop
                     /note="Rop protein, which maintains plasmids at low copy
                     number"
                     /translation="VTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     misc_feature    4960..5102
                     /label=bom
                     /note="basis of mobility region from pBR322"
     rep_origin      complement(5288..5876)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             5998..6810
                     /codon_start=1
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      complement(6906..7361)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtatgagg agggtagagg
      181 tgtctggagt ctctgtgatg gctgcctttc gggccgcttg gtcagccatc cggttgcctc
      241 tagcctcggc gctgtgtccc ttttgatgtc ctggacaatg gattatgcta agtcttttgg
      301 gcagaaagag ggcttttagt agggccaaga tctcgtcttt atttttgatc tctttgcctt
      361 ctgatgtgag caacccacgc cttctgtata tttctccatg gatatgggca gtagcaaaag
      421 cataacggct atcagtataa acatttagct tcttaccttc tgccatcttt agggcctggg
      481 tgagtgctat cagttcagcc cgctgagcgg atgtcccggc tggcagggct ttagcccaga
      541 ttacctcggt ctcggtggtc accgcagctc ccgccttacg ctgtccctct tgtaagagac
      601 tgcttccatc cgtgtaccag gtgtggtcgg cgtctgggag cggctggtcc gttaggtcgg
      661 gtcgggttcc gtgggcttcg gccaggatat caaggcagtt gtgttgcagc ccttcctcag
      721 gcagtgggag cagcgtagcc gggttcaggg ctaccaccgg tccgaactgg acccggtccg
      781 tgtccaaaag caaggcctga tagtgagtca tccgggcgtt ggaaagccag cggtcggggg
      841 gttgtttgac tagtgcctct actgcatggg gggccagaat gactagtggc tgtcccatgg
      901 ttagcttgcc tgcatccttt gtcagtacgg caatggctgc taccatccgt aggcaagggg
      961 gccacccagc tgctactggg tctagctttt tggacaggta ggccaccggc cgacgccaag
     1021 gtcccagttt ttgcgttagg acacctttgg cgtagccctg cttctcgtcg acaaagagtt
     1081 caaagggctt agtcaaatct ggcaacccca gggctggggc agttagaaga gcttgcttga
     1141 tttcttgata ggccttttgt tggtctgggc cccaattaaa cagagtcccc gttttggtga
     1201 gagggtacaa gggggctgcc atttctgcaa acccagggat ccagaggcga cagaagcctg
     1261 ccgtccctag gaactccctt agttgtcgag gggtcttcgg agtaggctgc cccatcacag
     1321 tctcttttct ggcctcagtc agccatctct gaccctcttt tagaagatac cccagatact
     1381 tgacctgttt ctggcaaatt tgggctttct tggccgaggc ccgatacccg aggttcccta
     1441 gggtttgtaa cagggcccga gtaccttgtt ggcagtctag ctcagaagtg gcggccagca
     1501 gtaagtcatc cacgtactgt agcaggatca agtctgggtg ctggatccgg aagtctgcta
     1561 ggtctctgtg cagtgcctca tcaaacaggg tgggactgtt tttgaaaccc tgtgggagtc
     1621 tggtccaggt caattgtcct gagattccca tctctggatc tctccactca aaggcgaaga
     1681 gaggctgact ggtggggtgg agtctcaggc agaaaaaggc atcctttaaa tcaagcacag
     1741 tgtaccactg gtgggacggt gggagcccgc tcaagaggtt gtaagggttg ggcacggtgg
     1801 ggtggatgtc ttccacccgc ttgttgactt ctctcagatc ctggacaggc ctataatcat
     1861 tagtccctgg tttcttaacg ggtagcaggg gcgtgttcca gggggactgg cagggtacca
     1921 gtattccctg gtccaacagt ctctgtatgt ggggcttgat ccccagtctg gcttcttgtg
     1981 acatggggta ttgttttatg gacacggggg tagaggttgc tttcagaggt atgatcagag
     2041 gagcttggcg aactgccagt cccatgcccc cggtttccgc ccaggcctga ggaaaatcag
     2101 acagccatgt ggaccctaga gaaacatctg gctcttttga ggtctcatgt agccgatgct
     2161 catcttctat atttagaagc ttgtcgacgg agctcgaatt cggatcccga cccatttgct
     2221 gtccaccagt catgctagcc atatggctgc cgcgcggcac caggccgctg ctgtgatgat
     2281 gatgatgatg gctgctgccc atggtatatc tccttcttaa agttaaacaa aattatttct
     2341 agaggggaat tgttatccgc tcacaattcc cctatagtga gtcgtattaa tttcgcggga
     2401 tcgagatctc gatcctctac gccggacgca tcgtggccgg catcaccggc gccacaggtg
     2461 cggttgctgg cgcctatatc gccgacatca ccgatgggga agatcgggct cgccacttcg
     2521 ggctcatgag cgcttgtttc ggcgtgggta tggtggcagg ccccgtggcc gggggactgt
     2581 tgggcgccat ctccttgcat gcaccattcc ttgcggcggc ggtgctcaac ggcctcaacc
     2641 tactactggg ctgcttccta atgcaggagt cgcataaggg agagcgtcga gatcccggac
     2701 accatcgaat ggcgcaaaac ctttcgcggt atggcatgat agcgcccgga agagagtcaa
     2761 ttcagggtgg tgaatgtgaa accagtaacg ttatacgatg tcgcagagta tgccggtgtc
     2821 tcttatcaga ccgtttcccg cgtggtgaac caggccagcc acgtttctgc gaaaacgcgg
     2881 gaaaaagtgg aagcggcgat ggcggagctg aattacattc ccaaccgcgt ggcacaacaa
     2941 ctggcgggca aacagtcgtt gctgattggc gttgccacct ccagtctggc cctgcacgcg
     3001 ccgtcgcaaa ttgtcgcggc gattaaatct cgcgccgatc aactgggtgc cagcgtggtg
     3061 gtgtcgatgg tagaacgaag cggcgtcgaa gcctgtaaag cggcggtgca caatcttctc
     3121 gcgcaacgcg tcagtgggct gatcattaac tatccgctgg atgaccagga tgccattgct
     3181 gtggaagctg cctgcactaa tgttccggcg ttatttcttg atgtctctga ccagacaccc
     3241 atcaacagta ttattttctc ccatgaagac ggtacgcgac tgggcgtgga gcatctggtc
     3301 gcattgggtc accagcaaat cgcgctgtta gcgggcccat taagttctgt ctcggcgcgt
     3361 ctgcgtctgg ctggctggca taaatatctc actcgcaatc aaattcagcc gatagcggaa
     3421 cgggaaggcg actggagtgc catgtccggt tttcaacaaa ccatgcaaat gctgaatgag
     3481 ggcatcgttc ccactgcgat gctggttgcc aacgatcaga tggcgctggg cgcaatgcgc
     3541 gccattaccg agtccgggct gcgcgttggt gcggatatct cggtagtggg atacgacgat
     3601 accgaagaca gctcatgtta tatcccgccg ttaaccacca tcaaacagga ttttcgcctg
     3661 ctggggcaaa ccagcgtgga ccgcttgctg caactctctc agggccaggc ggtgaagggc
     3721 aatcagctgt tgcccgtctc actggtgaaa agaaaaacca ccctggcgcc caatacgcaa
     3781 accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga
     3841 ctggaaagcg ggcagtgagc gcaacgcaat taatgtaagt tagctcactc attaggcacc
     3901 gggatctcga ccgatgccct tgagagcctt caacccagtc agctccttcc ggtgggcgcg
     3961 gggcatgact atcgtcgccg cacttatgac tgtcttcttt atcatgcaac tcgtaggaca
     4021 ggtgccggca gcgctctggg tcattttcgg cgaggaccgc tttcgctgga gcgcgacgat
     4081 gatcggcctg tcgcttgcgg tattcggaat cttgcacgcc ctcgctcaag ccttcgtcac
     4141 tggtcccgcc accaaacgtt tcggcgagaa gcaggccatt atcgccggca tggcggcccc
     4201 acgggtgcgc atgatcgtgc tcctgtcgtt gaggacccgg ctaggctggc ggggttgcct
     4261 tactggttag cagaatgaat caccgatacg cgagcgaacg tgaagcgact gctgctgcaa
     4321 aacgtctgcg acctgagcaa caacatgaat ggtcttcggt ttccgtgttt cgtaaagtct
     4381 ggaaacgcgg aagtcagcgc cctgcaccat tatgttccgg atctgcatcg caggatgctg
     4441 ctggctaccc tgtggaacac ctacatctgt attaacgaag cgctggcatt gaccctgagt
     4501 gatttttctc tggtcccgcc gcatccatac cgccagttgt ttaccctcac aacgttccag
     4561 taaccgggca tgttcatcat cagtaacccg tatcgtgagc atcctctctc gtttcatcgg
     4621 tatcattacc cccatgaaca gaaatccccc ttacacggag gcatcagtga ccaaacagga
     4681 aaaaaccgcc cttaacatgg cccgctttat cagaagccag acattaacgc ttctggagaa
     4741 actcaacgag ctggacgcgg atgaacaggc agacatctgt gaatcgcttc acgaccacgc
     4801 tgatgagctt taccgcagct gcctcgcgcg tttcggtgat gacggtgaaa acctctgaca
     4861 catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc
     4921 ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga cccagtcacg
     4981 tagcgatagc ggagtgtata ctggcttaac tatgcggcat cagagcagat tgtactgaga
     5041 gtgcaccata tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca
     5101 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     5161 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     5221 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     5281 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     5341 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     5401 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     5461 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     5521 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     5581 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     5641 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     5701 ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc
     5761 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     5821 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     5881 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     5941 ttttggtcat gaacaataaa actgtctgct tacataaaca gtaatacaag gggtgttatg
     6001 agccatattc aacgggaaac gtcttgctct aggccgcgat taaattccaa catggatgct
     6061 gatttatatg ggtataaatg ggctcgcgat aatgtcgggc aatcaggtgc gacaatctat
     6121 cgattgtatg ggaagcccga tgcgccagag ttgtttctga aacatggcaa aggtagcgtt
     6181 gccaatgatg ttacagatga gatggtcaga ctaaactggc tgacggaatt tatgcctctt
     6241 ccgaccatca agcattttat ccgtactcct gatgatgcat ggttactcac cactgcgatc
     6301 cccgggaaaa cagcattcca ggtattagaa gaatatcctg attcaggtga aaatattgtt
     6361 gatgcgctgg cagtgttcct gcgccggttg cattcgattc ctgtttgtaa ttgtcctttt
     6421 aacagcgatc gcgtatttcg tctcgctcag gcgcaatcac gaatgaataa cggtttggtt
     6481 gatgcgagtg attttgatga cgagcgtaat ggctggcctg ttgaacaagt ctggaaagaa
     6541 atgcataaac ttttgccatt ctcaccggat tcagtcgtca ctcatggtga tttctcactt
     6601 gataacctta tttttgacga ggggaaatta ataggttgta ttgatgttgg acgagtcgga
     6661 atcgcagacc gataccagga tcttgccatc ctatggaact gcctcggtga gttttctcct
     6721 tcattacaga aacggctttt tcaaaaatat ggtattgata atcctgatat gaataaattg
     6781 cagtttcatt tgatgctcga tgagtttttc taagaattaa ttcatgagcg gatacatatt
     6841 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     6901 acctgaaatt gtaaacgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
     6961 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
     7021 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
     7081 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
     7141 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
     7201 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
     7261 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
     7321 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc ca
//
