LOCUS       Exported                6729 bp DNA     circular SYN 10-SEP-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6729)
  AUTHORS   11111111
  TITLE     Direct Submission
  JOURNAL   Exported Sep 10, 2025 from SnapGene 6.0.2
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6729
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        191..209
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    210..234
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /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)."
     CDS             291..308
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             315..1799
                     /codon_start=1
                     /gene="nusA"
                     /product="transcription elongation protein (N utilization 
                     substance protein A)"
                     /label=NusA
                     /note="highly soluble in E. coli"
                     /translation="MNKEILAVVEAVSNEKALPREKIFEALESALATATKKKYEQEIDV
                     RVQIDRKSGDFDTFRRWLVVDEVTQPTKEITLEAARYEDESLNLGDYVEDQIESVTFDR
                     ITTQTAKQVIVQKVREAERAMVVDQFREHEGEIITGVVKKVNRDNISLDLGNNAEAVIL
                     REDMLPRENFRPGDRVRGVLYSVRPEARGAQLFVTRSKPEMLIELFRIEVPEIGEEVIE
                     IKAAARDPGSRAKIAVKTNDKRIDPVGACVGMRGARVQAVSTELGGERIDIVLWDDNPA
                     QFVINAMAPADVASIVVDEDKHTMDIAVEAGNLAQAIGRNGQNVRLASQLSGWELNVMT
                     VDDLQAKHQAEAHAAIDTFTKYLDIDEDFATVLVEEGFSTLEELAYVPMKELLEIEGLD
                     EPTVEALRERAKNALATIAQAQEESLGDNKPADDLLNLEGVDRDLAFKLAARGVCTLED
                     LAEQGIDDLADIEGLTDEKAGALIMAARNICWFGDEA"
     CDS             1815..1832
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1842..1865
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1944..1961
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1962..2006
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     terminator      2101..2148
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2185..2640
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     CDS             complement(2732..3547)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      3669..4257
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    4443..4582
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(4684..4875)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5447..6529)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(6530..6607)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 cccccggcca cggggcctgc caccataccc acgccgaaac aagcgctcat gagcccgaag
       61 tggcgagccc gatcttcccc atcggtgatg tcggcgatat aggcgccagc aaccgcacct
      121 gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga ggatcgagat cgatctcgat
      181 cccgcgaaat taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta
      241 gaaataattt tgtttaactt taagaaggag atatacatat gggcagcagc catcaccacc
      301 atcaccattc tagtatgaac aaagaaattt tggctgtagt tgaagccgta tccaatgaaa
      361 aggcgctacc tcgcgagaag attttcgaag cattggaaag cgcgctggcg acagcaacaa
      421 agaaaaaata tgaacaagag atcgacgtcc gcgtacagat cgatcgcaaa agcggtgatt
      481 ttgacacttt ccgtcgctgg ttagttgttg atgaagtcac ccagccgacc aaggaaatca
      541 cccttgaagc cgcacgttat gaagatgaaa gcctgaacct gggcgattac gttgaagatc
      601 agattgagtc tgttaccttt gaccgtatca ctacccagac ggcaaaacag gttatcgtgc
      661 agaaagtgcg tgaagccgaa cgtgcgatgg tggttgatca gttccgtgaa cacgaaggtg
      721 aaatcatcac cggcgtggtg aaaaaagtaa accgcgacaa catctctctg gatctgggca
      781 acaacgctga agccgtgatc ctgcgcgaag atatgctgcc gcgtgaaaac ttccgccctg
      841 gcgaccgcgt tcgtggcgtg ctctattccg ttcgcccgga agcgcgtggc gcgcaactgt
      901 tcgtcactcg ttccaagccg gaaatgctga tcgaactgtt ccgtattgaa gtgccagaaa
      961 tcggcgaaga agtgattgaa attaaagcag cggctcgcga tccgggttct cgtgcgaaaa
     1021 tcgcggtgaa aaccaacgat aaacgtatcg atccggtagg tgcttgcgta ggtatgcgtg
     1081 gcgcgcgtgt tcaggcggtg tctactgaac tgggtggcga gcgtatcgat atcgtcctgt
     1141 gggatgataa cccggcgcag ttcgtgatta acgcaatggc accggcagac gttgcttcta
     1201 tcgtggtgga tgaagataaa cacaccatgg acatcgccgt tgaagccggt aatctggcgc
     1261 aggcgattgg ccgtaacggt cagaacgtgc gtctggcttc gcaactgagc ggttgggaac
     1321 tcaacgtgat gaccgttgac gacctgcaag ctaagcatca ggcggaagcg cacgcagcga
     1381 tcgacacctt caccaaatat ctcgacatcg acgaagactt cgcgactgtt ctggtagaag
     1441 aaggcttctc gacgctggaa gaattggcct atgtgccgat gaaagagctg ttggaaatcg
     1501 aaggccttga tgagccgacc gttgaagcac tgcgcgagcg tgctaaaaat gcactggcca
     1561 ccattgcaca ggcccaggaa gaaagcctcg gtgataacaa accggctgac gatctgctga
     1621 accttgaagg ggtagatcgt gatttggcat tcaaactggc cgcccgtggc gtttgtacgc
     1681 tggaagatct cgccgaacag ggcattgatg atctggctga tatcgaaggg ttgaccgacg
     1741 aaaaagccgg agcactgatt atggctgccc gtaatatttg ctggttcggt gacgaagcga
     1801 ctagtggttc tggtcatcac catcaccatc actccgcggc tcttgaagtc ctctttcagg
     1861 gacccgggta ccaggatccg aattctgtac aggccttggc gcgcccgacg tccgtcgaca
     1921 agcttgcggc cgcagagctc gctctggtgc cacgcggtag taaagaaacc gctgctgcta
     1981 aattcgaacg ccagcacatg gacagctcta cttctgctgc tctcgaggct taattaacct
     2041 aggctgctaa acaaagcccg aaaggaagct gagttggctg ctgccaccgc tgagcaataa
     2101 ctagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgct gaaaggagga
     2161 actatatccg gattggcgaa tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     2221 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg
     2281 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     2341 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     2401 agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     2461 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta
     2521 tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa
     2581 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg cttacaattt
     2641 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     2701 ttcaaatatg tatccgctca tgaattaatt cttagaaaaa ctcatcgagc atcaaatgaa
     2761 actgcaattt attcatatca ggattatcaa taccatattt ttgaaaaagc cgtttctgta
     2821 atgaaggaga aaactcaccg aggcagttcc ataggatggc aagatcctgg tatcggtctg
     2881 cgattccgac tcgtccaaca tcaatacaac ctattaattt cccctcgtca aaaataaggt
     2941 tatcaagtga gaaatcacca tgagtgacga ctgaatccgg tgagaatggc aaaagtttat
     3001 gcatttcttt ccagacttgt tcaacaggcc agccattacg ctcgtcatca aaatcactcg
     3061 catcaaccaa accgttattc attcgtgatt gcgcctgagc gagacgaaat acgcgatcac
     3121 tgttaaaagg acaattacaa acaggaatcg aatgcaaccg gcgcaggaac actgccagcg
     3181 catcaacaat attttcacct gaatcaggat attcttctaa tacctggaat gctgttttgc
     3241 cggggatcgc agtggtgagt aaccatgcat catcaggagt acggataaaa tgcttgatgg
     3301 tcggaagagg cataaattcc gtcagccagt ttagtctgac catctcatct gtaacatcat
     3361 tggcaacgct acctttgcca tgtttcagaa acaactctgg cgcatcgggc ttcccataca
     3421 atcgatagat tgtcgcacct gattgcccga cattatcgcg agcccattta tacccatata
     3481 aatcagcatc catgttggaa tttaatcgcg gcctagagca agacgtttcc cgttgaatat
     3541 ggctcataac accccttgta ttactgttta tgtaagcaga cagttttatt gttcatgacc
     3601 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
     3661 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     3721 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     3781 actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc
     3841 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     3901 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     3961 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     4021 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     4081 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     4141 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     4201 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     4261 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc
     4321 tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat
     4381 accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag
     4441 cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg caatggtgca
     4501 ctctcagtac aatctgctct gatgccgcat agttaagcca gtatacactc cgctatcgct
     4561 acgtgactgg gtcatggctg cgccccgaca cccgccaaca cccgctgacg cgccctgacg
     4621 ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat
     4681 gtgtcagagg ttttcaccgt catcaccgaa acgcgcgagg cagctgcggt aaagctcatc
     4741 agcgtggtcg tgaagcgatt cacagatgtc tgcctgttca tccgcgtcca gctcgttgag
     4801 tttctccaga agcgttaatg tctggcttct gataaagcgg gccatgttaa gggcggtttt
     4861 ttcctgtttg gtcactgatg cctccgtgta agggggattt ctgttcatgg gggtaatgat
     4921 accgatgaaa cgagagagga tgctcacgat acgggttact gatgatgaac atgcccggtt
     4981 actggaacgt tgtgagggta aacaactggc ggtatggatg cggcgggacc agagaaaaat
     5041 cactcagggt caatgccagc gcttcgttaa tacagatgta ggtgttccac agggtagcca
     5101 gcagcatcct gcgatgcaga tccggaacat aatggtgcag ggcgctgact tccgcgtttc
     5161 cagactttac gaaacacgga aaccgaagac cattcatgtt gttgctcagg tcgcagacgt
     5221 tttgcagcag cagtcgcttc acgttcgctc gcgtatcggt gattcattct gctaaccagt
     5281 aaggcaaccc cgccagccta gccgggtcct caacgacagg agcacgatca tgctagtcat
     5341 gccccgcgcc caccggaagg agctgactgg gttgaaggct ctcaagggca tcggtcgaga
     5401 tcccggtgcc taatgagtga gctaacttac attaattgcg ttgcgctcac tgcccgcttt
     5461 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     5521 cggtttgcgt attgggcgcc agggtggttt ttcttttcac cagtgagacg ggcaacagct
     5581 gattgccctt caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc
     5641 ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg gatataacat gagctgtctt
     5701 cggtatcgtc gtatcccact accgagatgt ccgcaccaac gcgcagcccg gactcggtaa
     5761 tggcgcgcat tgcgcccagc gccatctgat cgttggcaac cagcatcgca gtgggaacga
     5821 tgccctcatt cagcatttgc atggtttgtt gaaaaccgga catggcactc cagtcgcctt
     5881 cccgttccgc tatcggctga atttgattgc gagtgagata tttatgccag ccagccagac
     5941 gcagacgcgc cgagacagaa cttaatgggc ccgctaacag cgcgatttgc tggtgaccca
     6001 atgcgaccag atgctccacg cccagtcgcg taccgtcttc atgggagaaa ataatactgt
     6061 tgatgggtgt ctggtcagag acatcaagaa ataacgccgg aacattagtg caggcagctt
     6121 ccacagcaat ggcatcctgg tcatccagcg gatagttaat gatcagccca ctgacgcgtt
     6181 gcgcgagaag attgtgcacc gccgctttac aggcttcgac gccgcttcgt tctaccatcg
     6241 acaccaccac gctggcaccc agttgatcgg cgcgagattt aatcgccgcg acaatttgcg
     6301 acggcgcgtg cagggccaga ctggaggtgg caacgccaat cagcaacgac tgtttgcccg
     6361 ccagttgttg tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc gcttccactt
     6421 tttcccgcgt tttcgcagaa acgtggctgg cctggttcac cacgcgggaa acggtctgat
     6481 aagagacacc ggcatactct gcgacatcgt ataacgttac tggtttcaca ttcaccaccc
     6541 tgaattgact ctcttccggg cgctatcatg ccataccgcg aaaggttttg cgccattcga
     6601 tggtgtccgg gatctcgacg ctctccctta tgcgactcct gcattaggaa gcagcccagt
     6661 agtaggttga ggccgttgag caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg
     6721 cccaacagt
//
