LOCUS       62056_20600        6183 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 6183)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6183
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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)."
     RBS             59..81
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             101..118
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     CDS             128..781
                     /codon_start=1
                     /label=GST
                     /note="glutathione S-transferase from Schistosoma
                     japonicum"
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     CDS             818..838
                     /codon_start=1
                     /label=TEV site
                     /note="tobacco etch virus (TEV) protease recognition and 
                     cleavage site"
                     /translation="ENLYFQG"
     CDS             907..924
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     terminator      991..1038
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1075..1530
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        1557..1661
                     /label=AmpR promoter
     CDS             1662..2519
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      2693..3281
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    complement(3467..3609)
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(3714..3902)
                     /codon_start=1
                     /label=rop
                     /note="Rop protein, which maintains plasmids at low copy
                     number"
                     /translation="VTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     protein_bind    complement(4677..4698)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(4714..5793)
                     /codon_start=1
                     /label=lacI
                     /note="lac repressor"
                     /translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(5794..5871)
                     /label=lacI promoter
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat gtctggttct catcatcatc atcatcatag
      121 cagcggtatg tcccctatac taggttattg gaaaattaag ggccttgtgc aacccactcg
      181 acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc gcgatgaagg
      241 tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc ttccttatta
      301 tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata tagctgacaa
      361 gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc ttgaaggagc
      421 ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact ttgaaactct
      481 caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag atcgtttatg
      541 tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt tgtatgacgc
      601 tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa aattagtttg
      661 ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat ccagcaagta
      721 tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc atcctccaaa
      781 atcggatctg ggccacacag gccatagatc tggtaccgag aacctatact tccagggtca
      841 taagggatcc tagccatggc tgatatcgaa ttcgagctcg tcgacaagct tgcggccgca
      901 ctcgagcacc accaccacca ccactgagat ccggctgcta acaaagcccg aaaggaagct
      961 gagttggctg ctgccaccgc tgagcaataa ctagcataac cccttggggc ctctaaacgg
     1021 gtcttgaggg gttttttgct gaaaggagga actatatccg gattggcgaa tgggacgcgc
     1081 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     1141 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     1201 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     1261 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc
     1321 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     1381 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     1441 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     1501 attttaacaa aatattaacg tttacaattt caggtggcac ttttcgggga aatgtgcgcg
     1561 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     1621 aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc
     1681 gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa
     1741 cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac
     1801 tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga
     1861 tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag
     1921 agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca
     1981 cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca
     2041 tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa
     2101 ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc
     2161 tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgcagca atggcaacaa
     2221 cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag
     2281 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct
     2341 ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac
     2401 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa
     2461 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt
     2521 aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat
     2581 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
     2641 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     2701 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     2761 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     2821 cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact
     2881 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     2941 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     3001 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     3061 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     3121 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     3181 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     3241 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     3301 ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc
     3361 ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc
     3421 gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg atgcggtatt
     3481 ttctccttac gcatctgtgc ggtatttcac accgcatata tggtgcactc tcagtacaat
     3541 ctgctctgat gccgcatagt taagccagta tacactccgc tatcgctacg tgactgggtc
     3601 atggctgcgc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     3661 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     3721 tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc gtggtcgtga
     3781 agcgattcac agatgtctgc ctgttcatcc gcgtccagct cgttgagttt ctccagaagc
     3841 gttaatgtct ggcttctgat aaagcgggcc atgttaaggg cggttttttc ctgtttggtc
     3901 actgatgcct ccgtgtaagg gggatttctg ttcatggggg taatgatacc gatgaaacga
     3961 gagaggatgc tcacgatacg ggttactgat gatgaacatg cccggttact ggaacgttgt
     4021 gagggtaaac aactggcggt atggatgcgg cgggaccaga gaaaaatcac tcagggtcaa
     4081 tgccagcgct tcgttaatac agatgtaggt gttccacagg gtagccagca gcatcctgcg
     4141 atgcagatcc ggaacataat ggtgcagggc gctgacttcc gcgtttccag actttacgaa
     4201 acacggaaac cgaagaccat tcatgttgtt gctcaggtcg cagacgtttt gcagcagcag
     4261 tcgcttcacg ttcgctcgcg tatcggtgat tcattctgct aaccagtaag gcaaccccgc
     4321 cagcctagcc gggtcctcaa cgacaggagc acgatcatgc gcacccgtgg ggccgccatg
     4381 ccggcgataa tggcctgctt ctcgccgaaa cgtttggtgg cgggaccagt gacgaaggct
     4441 tgagcgaggg cgtgcaagat tccgaatacc gcaagcgaca ggccgatcat cgtcgcgctc
     4501 cagcgaaagc ggtcctcgcc gaaaatgacc cagagcgctg ccggcacctg tcctacgagt
     4561 tgcatgataa agaagacagt cataagtgcg gcgacgatag tcatgccccg cgcccaccgg
     4621 aaggagctga ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg tgcctaatga
     4681 gtgagctaac ttacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     4741 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     4801 cgccagggtg gtttttcttt tcaccagtga gacgggcaac agctgattgc ccttcaccgc
     4861 ctggccctga gagagttgca gcaagcggtc cacgctggtt tgccccagca ggcgaaaatc
     4921 ctgtttgatg gtggttaacg gcgggatata acatgagctg tcttcggtat cgtcgtatcc
     4981 cactaccgag atgtccgcac caacgcgcag cccggactcg gtaatggcgc gcattgcgcc
     5041 cagcgccatc tgatcgttgg caaccagcat cgcagtggga acgatgccct cattcagcat
     5101 ttgcatggtt tgttgaaaac cggacatggc actccagtcg ccttcccgtt ccgctatcgg
     5161 ctgaatttga ttgcgagtga gatatttatg ccagccagcc agacgcagac gcgccgagac
     5221 agaacttaat gggcccgcta acagcgcgat ttgctggtga cccaatgcga ccagatgctc
     5281 cacgcccagt cgcgtaccgt cttcatggga gaaaataata ctgttgatgg gtgtctggtc
     5341 agagacatca agaaataacg ccggaacatt agtgcaggca gcttccacag caatggcatc
     5401 ctggtcatcc agcggatagt taatgatcag cccactgacg cgttgcgcga gaagattgtg
     5461 caccgccgct ttacaggctt cgacgccgct tcgttctacc atcgacacca ccacgctggc
     5521 acccagttga tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg cgtgcagggc
     5581 cagactggag gtggcaacgc caatcagcaa cgactgtttg cccgccagtt gttgtgccac
     5641 gcggttggga atgtaattca gctccgccat cgccgcttcc actttttccc gcgttttcgc
     5701 agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc tgataagaga caccggcata
     5761 ctctgcgaca tcgtataacg ttactggttt cacattcacc accctgaatt gactctcttc
     5821 cgggcgctat catgccatac cgcgaaaggt tttgcgccat tcgatggtgt ccgggatctc
     5881 gacgctctcc cttatgcgac tcctgcatta ggaagcagcc cagtagtagg ttgaggccgt
     5941 tgagcaccgc cgccgcaagg aatggtgcat gcaaggagat ggcgcccaac agtcccccgg
     6001 ccacggggcc tgccaccata cccacgccga aacaagcgct catgagcccg aagtggcgag
     6061 cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca cctgtggcgc
     6121 cggtgatgcc ggccacgatg cgtccggcgt agaggatcga gatcgatctc gatcccgcga
     6181 aat
//
