LOCUS       sequence-69799-e        4731 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 4731)
  TITLE     Series-13: Co-transformation vectors for Series-2 transfer vectors
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4731)
  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            "Unpublished"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..4731
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(10..1092)
                     /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     1054..1073
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1093..1170)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1216..1235
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1216..1234
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1365..1382
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1392..2492
                     /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     2466..2489
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             2547..2567
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     primer_bind     complement(2692..2710)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2706..2753
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(2983..3798)
                     /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"
     CDS             complement(2983..3798)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3706..3725
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(3799..3890)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3908..4543)
                     /direction=LEFT
                     /label=ColA ori
                     /note="Plasmids containing the ColA origin of replication 
                     can be propagated in E. coli cells that contain additional 
                     plasmids with compatible origins."
     primer_bind     complement(4053..4072)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
       61 atcggccaac gcgcggggag aggcggtttg cgtattgggc gccagggtgg tttttctttt
      121 caccagtgag acgggcaaca gctgattgcc cttcaccgcc tggccctgag agagttgcag
      181 caagcggtcc acgctggttt gccccagcag gcgaaaatcc tgtttgatgg tggttaacgg
      241 cgggatataa catgagctgt cttcggtatc gtcgtatccc actaccgaga tgtccgcacc
      301 aacgcgcagc ccggactcgg taatggcgcg cattgcgccc agcgccatct gatcgttggc
      361 aaccagcatc gcagtgggaa cgatgccctc attcagcatt tgcatggttt gttgaaaacc
      421 ggacatggca ctccagtcgc cttcccgttc cgctatcggc tgaatttgat tgcgagtgag
      481 atatttatgc cagccagcca gacgcagacg cgccgagaca gaacttaatg ggcccgctaa
      541 cagcgcgatt tgctggtgac ccaatgcgac cagatgctcc acgcccagtc gcgtaccgtc
      601 ttcatgggag aaaataatac tgttgatggg tgtctggtca gagacatcaa gaaataacgc
      661 cggaacatta gtgcaggcag cttccacagc aatggcatcc tggtcatcca gcggatagtt
      721 aatgatcagc ccactgacgc gttgcgcgag aagattgtgc accgccgctt tacaggcttc
      781 gacgccgctt cgttctacca tcgacaccac cacgctggca cccagttgat cggcgcgaga
      841 tttaatcgcc gcgacaattt gcgacggcgc gtgcagggcc agactggagg tggcaacgcc
      901 aatcagcaac gactgtttgc ccgccagttg ttgtgccacg cggttgggaa tgtaattcag
      961 ctccgccatc gccgcttcca ctttttcccg cgttttcgca gaaacgtggc tggcctggtt
     1021 caccacgcgg gaaacggtct gataagagac accggcatac tctgcgacat cgtataacgt
     1081 tactggtttc acattcacca ccctgaattg actctcttcc gggcgctatc atgccatacc
     1141 gcgaaaggtt ttgcgccatt cgatggtgtc cgggatctcg acgctctccc ttatgcgact
     1201 cctgcattag gaaattaata cgactcacta tagggagacc acaacggttt ccctctagtg
     1261 ccggctccgg agagctcttt aattaagcgg ccgccctgca ggactcgagt tctagaaata
     1321 attttgttta actttaataa ggagatatac atatgaaatc ttctcaccat caccatcacc
     1381 atggttcttc tatgaaaatc gaagaaggta aactggtaat ctggattaac ggcgataaag
     1441 gctataacgg tctcgctgaa gtcggtaaga aattcgagaa agataccgga attaaagtca
     1501 ccgttgagca tccggataaa ctggaagaga aattcccaca ggttgcggca actggcgatg
     1561 gccctgacat tatcttctgg gcacacgacc gctttggtgg ctacgctcaa tctggcctgt
     1621 tggctgaaat caccccggac aaagcgttcc aggacaagct gtatccgttt acctgggatg
     1681 ccgtacgtta caacggcaag ctgattgctt acccgatcgc tgttgaagcg ttatcgctga
     1741 tttataacaa agatctgctg ccgaacccgc caaaaacctg ggaagagatc ccggcgctgg
     1801 ataaagaact gaaagcgaaa ggtaagagcg cgctgatgtt caacctgcaa gaaccgtact
     1861 tcacctggcc gctgattgct gctgacgggg gttatgcgtt caagtatgaa aacggcaagt
     1921 acgacattaa agacgtgggc gtggataacg ctggcgcgaa agcgggtctg accttcctgg
     1981 ttgacctgat taaaaacaaa cacatgaatg cagacaccga ttactccatc gcagaagctg
     2041 cctttaataa aggcgaaaca gcgatgacca tcaacggccc gtgggcatgg tccaacatcg
     2101 acaccagcaa agtgaattat ggtgtaacgg tactgccgac cttcaagggt caaccatcca
     2161 aaccgttcgt tggcgtgctg agcgcaggta ttaacgccgc cagtccgaac aaagagctgg
     2221 caaaagagtt cctcgaaaac tatctgctga ctgatgaagg tctggaagcg gttaataaag
     2281 acaaaccgct gggtgccgta gcgctgaagt cttacgagga agagttggcg aaagatccac
     2341 gtattgccgc cactatggaa aacgcccaga aaggtgaaat catgccgaac atcccgcaga
     2401 tgtccgcttt ctggtatgcc gtgcgtactg cggtgatcaa cgccgccagc ggtcgtcaga
     2461 ctgtcgatga agccctgaaa gacgcgcaga ctaattcgag ctcgaacaac aacaacaata
     2521 acaataacaa caacctcggg atcgaggaaa acctgtactt ccaatccaat attggaagtg
     2581 gataacggat ccgcgatcgc ggcgcgccac ctggtggccg gccggtacca cgcgtgcgcg
     2641 ctgatccggc tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc accgctgagc
     2701 aataactagc ataacccctt ggggcctcta aacgggtctt gaggggtttt ttgctgaaac
     2761 ctcaggcatt tgagaagcac acggtcacac tgcttccggt agtcaataaa ccggtaaacc
     2821 agcaatagac ataagcggct atttaacgac cctgccctga accgacgaca agctgacgac
     2881 cgggtctccg caagtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt
     2941 ttctaaatac attcaaatat gtatccgctc atgaattaat tcttagaaaa actcatcgag
     3001 catcaaatga aactgcaatt tattcatatc aggattatca ataccatatt tttgaaaaag
     3061 ccgtttctgt aatgaaggag aaaactcacc gaggcagttc cataggatgg caagatcctg
     3121 gtatcggtct gcgattccga ctcgtccaac atcaatacaa cctattaatt tcccctcgtc
     3181 aaaaataagg ttatcaagtg agaaatcacc atgagtgacg actgaatccg gtgagaatgg
     3241 caaaagttta tgcatttctt tccagacttg ttcaacaggc cagccattac gctcgtcatc
     3301 aaaatcactc gcatcaacca aaccgttatt cattcgtgat tgcgcctgag cgagacgaaa
     3361 tacgcggtcg ctgttaaaag gacaattaca aacaggaatc gaatgcaacc ggcgcaggaa
     3421 cactgccagc gcatcaacta tattttcacc tgaatcagga tattcttcta atacctggaa
     3481 tgctgttttc ccggggatcg cagtggtgag taaccatgca tcatcaggag tacggataaa
     3541 atgcttgatg gtcggaagag gcataaattc cgtcagccag tttagtctga ccatctcatc
     3601 tgtaacatca ttggcaacgc tacctttgcc atgtttcaga aacaactctg gcgcatcggg
     3661 cttcccatac aatcgataga ttgtcgcacc tgattgcccg acattatcgc gagcccattt
     3721 atacccatat aaatcagcat ccatgttgga atttaatcgc ggcctagagc aagacgtttc
     3781 ccgttgaata tggctcatac tcttcctttt tcattattat tgaagcattt atcagggtta
     3841 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggcatgct
     3901 agcgcagaaa cgtcctagaa gatgccagga ggatacttag cagagagaca ataaggccgg
     3961 agcgaagccg tttttccata ggctccgccc ccctgacgaa catcacgaaa tctgacgctc
     4021 aaatcagtgg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctgatgg
     4081 ctccctcttg cgctctcctg ttcccgtcct gcggcgtccg tgttgtggtg gaggctttac
     4141 ccaaatcacc acgtcccgtt ccgtgtagac agttcgctcc aagctgggct gtgtgcaaga
     4201 accccccgtt cagcccgact gctgcgcctt atccggtaac tatcatcttg agtccaaccc
     4261 ggaaagacac gacaaaacgc cactggcagc agccattggt aactgagaat tagtggattt
     4321 agttatcgag agtcttgaag tggtggccta acagaggcta cactgaaagg acagtatttg
     4381 gtatctgcgc tccactaaag ccagttacca ggttaagcag ttccccaact gacttaacct
     4441 tcgatcaaac cgcctcccca ggcggttttt tcgtttacag agcaggagat tacgacgatc
     4501 gtaaaaggat ctcaagaaga tcctttacgg attcccgaca ccatcactct tgatttcagt
     4561 gcaatttatc tcttcaaatg tagcacctga agtcagcccc atacgatata agttgtaatt
     4621 ctcatgttag tcatgccccg cgcccaccgg aaggagctga ctgggttgaa ggctctcaag
     4681 ggcatcggtc gagatcccgg tgcctaatga gtgagctaac ttacattaat t
//
