LOCUS       Exported                6676 bp DNA     circular SYN 22-AUG-2020
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6676)
  AUTHORS   Youbio
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene 6.0.2
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6676
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        3..80
                     /gene="lacI (mutant)"
                     /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             81..1163
                     /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"
     misc_feature    1406..1434
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1442..1458
                     /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             1531..1548
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1558..2655
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="KTEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLEE
                     KFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKLI
                     AYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIAA
                     DGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGET
                     AMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFLE
                     NYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAFW
                     YAVRTAVINAASGRQTVDEALKDAQT"
     CDS             2704..2718
                     /codon_start=1
                     /product="enterokinase recognition and cleavage site"
                     /label=enterokinase site
                     /translation="DDDDK"
     primer_bind     complement(2766..2782)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rRNA            3011..3130
                     /product="5S ribosomal RNA"
                     /label=5S ribosomal RNA
                     /note="The 5S rRNA gene is from the E. coli rrnB operon"
     terminator      3131..3336
                     /label=rrnB T1/T2 terminator
     promoter        3356..3447
                     /gene="bla"
                     /label=AmpR promoter
     CDS             3448..4308
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERSPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      complement(4408..4863)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     rep_origin      4974..5562
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    5748..5890
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(5992..6183)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 ccgacaccat cgaatggtgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga
       61 gtcaattcag ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg
      121 gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa
      181 cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac
      241 aacaactggc gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc
      301 acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg
      361 tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc
      421 ttctcgcgca acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca
      481 ttgctgtgga agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga
      541 cacccatcaa cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc
      601 tggtcgcatt gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg
      661 cgcgtctgcg tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag
      721 cggaacggga aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga
      781 atgagggcat cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa
      841 tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga tatctcggta gtgggatacg
      901 acgataccga agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc
      961 gcctgctggg gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga
     1021 agggcaatca gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata
     1081 cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt
     1141 cccgactgga aagcgggcag tgagcgcaac gcaattaatg taagttagct cactcattag
     1201 gcacaattct catgtttgac agcttatcat cgactgcacg gtgcaccaat gcttctggcg
     1261 tcaggcagcc atcggaagct gtggtatggc tgtgcaggtc gtaaatcact gcataattcg
     1321 tgtcgctcaa ggcgcactcc cgttctggat aatgtttttt gcgccgacat cataacggtt
     1381 ctggcaaata ttctgaaatg agctgttgac aattaatcat cggctcgtat aatgtgtgga
     1441 attgtgagcg gataacaatt tcacacagga aacagccagt ccgtttaggt gttttcacga
     1501 gcacttcacc aacaaggacc atagcatatg catcatcacc atcaccatag cagcggcaaa
     1561 actgaagaag gtaaactggt aatctggatt aacggcgata aaggctataa cggtctcgct
     1621 gaagtcggta agaaattcga gaaagatacc ggaattaaag tcaccgttga gcatccggat
     1681 aaactggaag agaaattccc acaggttgcg gcaactggcg atggccctga cattatcttc
     1741 tgggcacacg accgctttgg tggctacgct caatctggcc tgttggctga aatcaccccg
     1801 gacaaagcgt tccaggacaa gctgtatccg tttacctggg atgccgtacg ttacaacggc
     1861 aagctgattg cttacccgat cgctgttgaa gcgttatcgc tgatttataa caaagatctg
     1921 ctgccgaacc cgccaaaaac ctgggaagag atcccggcgc tggataaaga actgaaagcg
     1981 aaaggtaaga gcgcgctgat gttcaacctg caagaaccgt acttcacctg gccgctgatt
     2041 gctgctgacg ggggttatgc gttcaagtat gaaaacggca agtacgacat taaagacgtg
     2101 ggcgtggata acgctggcgc gaaagcgggt ctgaccttcc tggttgacct gattaaaaac
     2161 aaacacatga atgcagacac cgattactcc atcgcagaag ctgcctttaa taaaggcgaa
     2221 acagcgatga ccatcaacgg cccgtgggca tggtccaaca tcgacaccag caaagtgaat
     2281 tatggtgtaa cggtactgcc gaccttcaag ggtcaaccat ccaaaccgtt cgttggcgtg
     2341 ctgagcgcag gtattaacgc cgccagtccg aacaaagagc tggcaaaaga gttcctcgaa
     2401 aactatctgc tgactgatga aggtctggaa gcggttaata aagacaaacc gctgggtgcc
     2461 gtagcgctga agtcttacga ggaagagttg gcgaaagatc cacgtattgc cgccaccatg
     2521 gaaaacgccc agaaaggtga aatcatgccg aacatcccgc agatgtccgc tttctggtat
     2581 gccgtgcgta ctgcggtgat caacgccgcc agcggtcgtc agactgtcga tgaagccctg
     2641 aaagacgcgc agactaattc gagctcgaac aacaacaaca ataacaataa caacaacctc
     2701 ggggatgacg atgacaaggt accggaattc ggatcctcta gagtcgacct gcaggcaagc
     2761 ttggcactgg ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt
     2821 aatcgccttg cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc
     2881 gatcgccctt cccaacagtt gcgcagcctg aatggcgaat ggcagcttgg ctgttttggc
     2941 ggatgagata agattttcag cctgatacag attaaatcag aacgcagaag cggtctgata
     3001 aaacagaatt tgcctggcgg cagtagcgcg gtggtcccac ctgaccccat gccgaactca
     3061 gaagtgaaac gccgtagcgc cgatggtagt gtggggtctc cccatgcgag agtagggaac
     3121 tgccaggcat caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg
     3181 ttgtttgtcg gtgaacgctc tcctgagtag gacaaatccg ccgggagcgg atttgaacgt
     3241 tgcgaagcaa cggcccggag ggtggcgggc aggacgcccg ccataaactg ccaggcatca
     3301 aattaagcag aaggccatcc tgacggatgg cctttttgcg tttctacaaa ctctttttgt
     3361 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     3421 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     3481 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     3541 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     3601 ggtaagatcc ttgagagttt tcgccccgaa gaacgttctc caatgatgag cacttttaaa
     3661 gttctgctat gtggcgcggt attatcccgt gttgacgccg ggcaagagca actcggtcgc
     3721 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     3781 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     3841 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     3901 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     3961 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
     4021 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     4081 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     4141 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     4201 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     4261 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     4321 gtttactcat atatacttta gattgattta ccccggttga taatcagaaa agccccaaaa
     4381 acaggaagat tgtataagca aatatttaaa ttgtaaacgt taatattttg ttaaaattcg
     4441 cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc
     4501 cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga
     4561 gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg
     4621 atggcccact acgtgaacca tcacccaaat caagtttttt ggggtcgagg tgccgtaaag
     4681 cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga
     4741 acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg
     4801 tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg
     4861 cgtaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     4921 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     4981 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     5041 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     5101 gcgcagatac caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac
     5161 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     5221 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     5281 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     5341 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     5401 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     5461 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     5521 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     5581 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     5641 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     5701 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgcct gatgcggtat
     5761 tttctcctta cgcatctgtg cggtatttca caccgcatat atggtgcact ctcagtacaa
     5821 tctgctctga tgccgcatag ttaagccagt atacactccg ctatcgctac gtgactgggt
     5881 catggctgcg ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct
     5941 cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt
     6001 ttcaccgtca tcaccgaaac gcgcgaggca gctgcggtaa agctcatcag cgtggtcgtg
     6061 cagcgattca cagatgtctg cctgttcatc cgcgtccagc tcgttgagtt tctccagaag
     6121 cgttaatgtc tggcttctga taaagcgggc catgttaagg gcggtttttt cctgtttggt
     6181 cactgatgcc tccgtgtaag ggggatttct gttcatgggg gtaatgatac cgatgaaacg
     6241 agagaggatg ctcacgatac gggttactga tgatgaacat gcccggttac tggaacgttg
     6301 tgagggtaaa caactggcgg tatggatgcg gcgggaccag agaaaaatca ctcagggtca
     6361 atgccagcgc ttcgttaata cagatgtagg tgttccacag ggtagccagc agcatcctgc
     6421 gatgcagatc cggaacataa tggtgcaggg cgctgacttc cgcgtttcca gactttacga
     6481 aacacggaaa ccgaagacca ttcatgttgt tgctcaggtc gcagacgttt tgcagcagca
     6541 gtcgcttcac gttcgctcgc gtatcggtga ttcattctgc taaccagtaa ggcaaccccg
     6601 ccagcctagc cgggtcctca acgacaggag cacgatcatg cgcacccgtg gccaggaccc
     6661 aacgctgccc gaaatt
//
