LOCUS       sequence-69807-e        4785 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4785)
  TITLE     Series-13: Co-transformation vectors for Series-2 transfer vectors
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4785)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pET His6 MBP TEV co-transformation cloning vec...
FEATURES             Location/Qualifiers
     source          1..4785
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(3..1085)
                     /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"
     primer_bind     1047..1066
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1086..1163)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1209..1228
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1209..1227
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1232..1255
                     /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             1350..1367
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1377..2477
                     /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="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     2451..2474
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             2532..2552
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     primer_bind     complement(2677..2695)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2691..2738
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(2909..3700)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     promoter        complement(3701..3792)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3840..4578)
                     /direction=LEFT
                     /label=CloDF13 ori
                     /note="Plasmids containing the CloDF13 (CDF) origin of 
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
ORIGIN
        1 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
       61 aacgcgcggg gagaggcggt ttgcgtattg ggcgccaggg tggtttttct tttcaccagt
      121 gagacgggca acagctgatt gcccttcacc gcctggccct gagagagttg cagcaagcgg
      181 tccacgctgg tttgccccag caggcgaaaa tcctgtttga tggtggttaa cggcgggata
      241 taacatgagc tgtcttcggt atcgtcgtat cccactaccg agatgtccgc accaacgcgc
      301 agcccggact cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc
      361 atcgcagtgg gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa accggacatg
      421 gcactccagt cgccttcccg ttccgctatc ggctgaattt gattgcgagt gagatattta
      481 tgccagccag ccagacgcag acgcgccgag acagaactta atgggcccgc taacagcgcg
      541 atttgctggt gacccaatgc gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg
      601 gagaaaataa tactgttgat gggtgtctgg tcagagacat caagaaataa cgccggaaca
      661 ttagtgcagg cagcttccac agcaatggca tcctggtcat ccagcggata gttaatgatc
      721 agcccactga cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc ttcgacgccg
      781 cttcgttcta ccatcgacac caccacgctg gcacccagtt gatcggcgcg agatttaatc
      841 gccgcgacaa tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac gccaatcagc
      901 aacgactgtt tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc
      961 atcgccgctt ccactttttc ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg
     1021 cgggaaacgg tctgataaga gacaccggca tactctgcga catcgtataa cgttactggt
     1081 ttcacattca ccaccctgaa ttgactctct tccgggcgct atcatgccat accgcgaaag
     1141 gttttgcgcc attcgatggt gtccgggatc tcgacgctct cccttatgcg actcctgcat
     1201 taggaaatta atacgactca ctatagggag agaattgtga gcggataaca attcccctgt
     1261 agattaatta agcggccgcc ctgcaggact cgagttctag aaataatttt gtttaacttt
     1321 aataaggaga tatacatatg aaatcttctc accatcacca tcaccatggt tcttctatga
     1381 aaatcgaaga aggtaaactg gtaatctgga ttaacggcga taaaggctat aacggtctcg
     1441 ctgaagtcgg taagaaattc gagaaagata ccggaattaa agtcaccgtt gagcatccgg
     1501 ataaactgga agagaaattc ccacaggttg cggcaactgg cgatggccct gacattatct
     1561 tctgggcaca cgaccgcttt ggtggctacg ctcaatctgg cctgttggct gaaatcaccc
     1621 cggacaaagc gttccaggac aagctgtatc cgtttacctg ggatgccgta cgttacaacg
     1681 gcaagctgat tgcttacccg atcgctgttg aagcgttatc gctgatttat aacaaagatc
     1741 tgctgccgaa cccgccaaaa acctgggaag agatcccggc gctggataaa gaactgaaag
     1801 cgaaaggtaa gagcgcgctg atgttcaacc tgcaagaacc gtacttcacc tggccgctga
     1861 ttgctgctga cgggggttat gcgttcaagt atgaaaacgg caagtacgac attaaagacg
     1921 tgggcgtgga taacgctggc gcgaaagcgg gtctgacctt cctggttgac ctgattaaaa
     1981 acaaacacat gaatgcagac accgattact ccatcgcaga agctgccttt aataaaggcg
     2041 aaacagcgat gaccatcaac ggcccgtggg catggtccaa catcgacacc agcaaagtga
     2101 attatggtgt aacggtactg ccgaccttca agggtcaacc atccaaaccg ttcgttggcg
     2161 tgctgagcgc aggtattaac gccgccagtc cgaacaaaga gctggcaaaa gagttcctcg
     2221 aaaactatct gctgactgat gaaggtctgg aagcggttaa taaagacaaa ccgctgggtg
     2281 ccgtagcgct gaagtcttac gaggaagagt tggcgaaaga tccacgtatt gccgccacta
     2341 tggaaaacgc ccagaaaggt gaaatcatgc cgaacatccc gcagatgtcc gctttctggt
     2401 atgccgtgcg tactgcggtg atcaacgccg ccagcggtcg tcagactgtc gatgaagccc
     2461 tgaaagacgc gcagactaat tcgagctcga acaacaacaa caataacaat aacaacaacc
     2521 tcgggatcga ggaaaacctg tacttccaat ccaatattgg aagtggataa cggatccgcg
     2581 atcgcggcgc gccacctggt ggccggccgg taccacgcgt gcgcgctgat ccggctgcta
     2641 acaaagcccg aaaggaagct gagttggctg ctgccaccgc tgagcaataa ctagcataac
     2701 cccttggggc ctctaaacgg gtcttgaggg gttttttgct gaaacctcag gcatttgaga
     2761 agcacacggt cacactgctt ccggtagtca ataaaccggt aaaccagcaa tagacataag
     2821 cggctattta acgaccctgc cctgaaccga cgaccgggtc atcgtggccg gatcttgcgg
     2881 cccctcggct tgaacgaatt gttagacatt atttgccgac taccttggtg atctcgcctt
     2941 tcacgtaatg gacaaattct tccaactgat ctgcgcgcga ggccaagcga tcttcttctt
     3001 gtccaagata agcctgtcta gcttcaagta tgacgggctg atactgggcc ggcaggcgct
     3061 ccattgccca gtcggcagcg acatccttcg gcgcgatttt gccggttact gcgctgtacc
     3121 aaatgcggga caacgtaagc actacatttc gctcatcgcc agcccagtcg ggcggcgagt
     3181 tccatagcgt taaggtttca tttagcgcct caaatagatc ctgttcagga accggatcaa
     3241 agagttcctc cgccgctgga cctaccaagg caacgctatg ttctcttgct tttgtcagca
     3301 agatagccag atcaatgtcg atcgtggctg gctcgaagat acctgcaaga atgtcattgc
     3361 gctgccattc tccaaattgc agttcgcgct tagctggata acgccacgga atgatgtcgt
     3421 cgtgcacaac aatggtgact tctacagcgc ggagaatctc gctctctcca ggggaagccg
     3481 aagtttccaa aaggtcgttg atcaaagctc gccgcgttgt ttcatcaagc cttacggtca
     3541 ccgtaaccag caaatcaata tcactgtgcg gcttcaggcc gccatccact gcggagccgt
     3601 acaaatgtac ggccagcaac gtcggttcga gatggcgctc gatgacgcca actacctctg
     3661 atagttgagt cgatacttcg gcgatcaccg cttccctcat actcttcctt tttcattatt
     3721 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     3781 aaaataaaca aatagctagc tcactcggtc gctacgctcc gggcgtgaga ctgcggcggg
     3841 cgctgcggac acatacaaag ttacccacag attccgtgga taagcagggg actaacatgt
     3901 gaggcaaaac agcagggccg cgccggtggc gtttttccat aggctccgcc ctcctgccag
     3961 agttcacata aacagacgct tttccggtgc atctgtggga gccgtgaggc tcaaccatga
     4021 atctgacagt acgggcgaaa cccgacagga cttaaagatc cccaccgttt ccggcgggtc
     4081 gctccctctt gcgctctcct gttccgaccc tgccgtttac cggatacctg ttccgccttt
     4141 ctcccttacg ggaagtgtgg cgctttctca tagctcacac actggtatct cggctcggtg
     4201 taggtcgttc gctccaagct gggctgtaag caagaactcc ccgttcagcc cgactgctgc
     4261 gccttatccg gtaactgttc acttgagtcc aacccggaaa agcacggtaa aacgccactg
     4321 gcagcagcca ttggtaactg ggagttcgca gaggatttgt ttagctaaac acgcggttgc
     4381 tcttgaagtg tgcgccaaag tccggctaca ctggaaggac agatttggtt gctgtgctct
     4441 gcgaaagcca gttaccacgg ttaagcagtt ccccaactga cttaaccttc gatcaaacca
     4501 cctccccagg tggttttttc gtttacaggg caaaagatta cgcgcagaaa aaaaggatct
     4561 caagaagatc ctttgatctt ttctactgaa ccgctcttga tttcagtgca atttatctct
     4621 tcaaatgtag cacctgaagt cagccccata cgatataagt tgtaattctc atgttagtca
     4681 tgccccgcgc ccaccggaag gagctgactg ggttgaaggc tctcaagggc atcggtcgag
     4741 atcccggtgc ctaatgagtg agctaactta cattaattgc gttgc
//
