LOCUS       sequence-7239-ex        4298 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 4298)
  AUTHORS   Iwai H, Zuger S, Jin J, Tam PH
  TITLE     Highly efficient protein trans-splicing by a naturally split DnaE 
            intein from Nostoc punctiforme.
  JOURNAL   FEBS Lett. 2006 Mar 20. 580(7):1853-8.
  PUBMED    16516207
REFERENCE   2  (bases 1 to 4298)
  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: "FEBS Lett. 
            2006 Mar 20. 580(7):1853-8."
COMMENT     Sequence Label: pSKDuet01
FEATURES             Location/Qualifiers
     source          1..4298
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        30..48
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    49..73
                     /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             129..146
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             156..173
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     promoter        729..747
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    748..772
                     /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             881..925
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(963..981)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      977..1024
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1254..2069)
                     /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(1254..2069)
                     /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     1977..1996
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(2070..2161)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2177..2926)
                     /direction=LEFT
                     /label=RSF ori
                     /note="Plasmids containing the RSF 1030 origin of 
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
     primer_bind     complement(2187..2204)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(2427..2446)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3122..4204)
                     /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     4166..4185
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4205..4282)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 tcccttatgc gactcctgca ttaggaaatt aatacgactc actatagggg aattgtgagc
       61 ggataacaat tcccctgtag aaataatttt gtttaacttt aataaggaga tataccatgg
      121 gcagcagcca tcatcatcat catcacagca gcggcctggt gccgcgcggc agccatatgc
      181 agtacaaact tatcctgaac ggtaaaaccc tgaaaggtga aaccaccacc gaagctgttg
      241 acgctgctac cgcggaaaaa gttttcaaac agtacgctaa cgacaacggt gttgacggtg
      301 aatggaccta cgacgacgct accaaaacct acaccgtaac ggaaggatcc tgtttaagct
      361 atgaaacgga aatattgaca gtagaatatg gattattacc gattggtaaa attgtagaaa
      421 agcgcatcga atgtactgtt tatagcgttg ataataatgg aaatatttat acacaacctg
      481 tagcacaatg gcacgatcgc ggagaacaag aggtgtttga gtattgtttg gaagatggtt
      541 cattgattcg ggcaacaaaa gaccataagt ttatgactgt tgatggtcaa atgttgccaa
      601 ttgatgaaat atttgaacgt gaattggatt tgatgcgggt tgataatttg ccgaattaag
      661 cttgcggccg cataatgctt aagtcgaaca gaaagtaatc gtattgtaca cggccgcata
      721 atcgaaatta atacgactca ctatagggga attgtgagcg gataacaatt ccccatctta
      781 gtatattagt taagtataag aaggagatat acatatggca gatctcaatt ggatatcggc
      841 cggccacgcg atcgctgacg tcggtaccct cgagtctggt aaagaaaccg ctgctgcgaa
      901 atttgaacgc cagcacatgg actcgtctac tagcgcagct taattaacct aggctgctgc
      961 caccgctgag caataactag cataacccct tggggcctct aaacgggtct tgaggggttt
     1021 tttgctgaaa cctcaggcat ttgagaagca cacggtcaca ctgcttccgg tagtcaataa
     1081 accggtaaac cagcaataga cataagcggc tatttaacga ccctgccctg aaccgacgac
     1141 aagctgacga ccgggtctcc gcaagtggca cttttcgggg aaatgtgcgc ggaaccccta
     1201 tttgtttatt tttctaaata cattcaaata tgtatccgct catgaattaa ttcttagaaa
     1261 aactcatcga gcatcaaatg aaactgcaat ttattcatat caggattatc aataccatat
     1321 ttttgaaaaa gccgtttctg taatgaagga gaaaactcac cgaggcagtt ccataggatg
     1381 gcaagatcct ggtatcggtc tgcgattccg actcgtccaa catcaataca acctattaat
     1441 ttcccctcgt caaaaataag gttatcaagt gagaaatcac catgagtgac gactgaatcc
     1501 ggtgagaatg gcaaaagttt atgcatttct ttccagactt gttcaacagg ccagccatta
     1561 cgctcgtcat caaaatcact cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga
     1621 gcgagacgaa atacgcggtc gctgttaaaa ggacaattac aaacaggaat cgaatgcaac
     1681 cggcgcagga acactgccag cgcatcaaca atattttcac ctgaatcagg atattcttct
     1741 aatacctgga atgctgtttt cccggggatc gcagtggtga gtaaccatgc atcatcagga
     1801 gtacggataa aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg
     1861 accatctcat ctgtaacatc attggcaacg ctacctttgc catgtttcag aaacaactct
     1921 ggcgcatcgg gcttcccata caatcgatag attgtcgcac ctgattgccc gacattatcg
     1981 cgagcccatt tatacccata taaatcagca tccatgttgg aatttaatcg cggcctagag
     2041 caagacgttt cccgttgaat atggctcata ctcttccttt ttcaatatta ttgaagcatt
     2101 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     2161 ataggcatgc agcgctcttc cgcttcctcg ctcactgact cgctacgctc ggtcgttcga
     2221 ctgcggcgag cggtgtcagc tcactcaaaa gcggtaatac ggttatccac agaatcaggg
     2281 gataaagccg gaaagaacat gtgagcaaaa agcaaagcac cggaagaagc caacgccgca
     2341 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagcca
     2401 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     2461 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     2521 gggaagcgtg gcgctttctc atagctcacg ctgttggtat ctcagttcgg tgtaggtcgt
     2581 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     2641 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     2701 cattggtaac tgatttagag gactttgtct tgaagttatg cacctgttaa ggctaaactg
     2761 aaagaacaga ttttggtgag tgcggtcctc caacccactt accttggttc aaagagttgg
     2821 tagctcagcg aaccttgaga aaaccaccgt tggtagcggt ggtttttctt tatttatgag
     2881 atgatgaatc aatcggtcta tcaagtcaac gaacagctat tccgttactc tagatttcag
     2941 tgcaatttat ctcttcaaat gtagcacctg aagtcagccc catacgatat aagttgtaat
     3001 tctcatgtta gtcatgcccc gcgcccaccg gaaggagctg actgggttga aggctctcaa
     3061 gggcatcggt cgagatcccg gtgcctaatg agtgagctaa cttacattaa ttgcgttgcg
     3121 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     3181 acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg
     3241 agacgggcaa cagctgattg cccttcaccg cctggccctg agagagttgc agcaagcggt
     3301 ccacgctggt ttgccccagc aggcgaaaat cctgtttgat ggtggttaac ggcgggatat
     3361 aacatgagct gtcttcggta tcgtcgtatc ccactaccga gatgtccgca ccaacgcgca
     3421 gcccggactc ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca
     3481 tcgcagtggg aacgatgccc tcattcagca tttgcatggt ttgttgaaaa ccggacatgg
     3541 cactccagtc gccttcccgt tccgctatcg gctgaatttg attgcgagtg agatatttat
     3601 gccagccagc cagacgcaga cgcgccgaga cagaacttaa tgggcccgct aacagcgcga
     3661 tttgctggtg acccaatgcg accagatgct ccacgcccag tcgcgtaccg tcttcatggg
     3721 agaaaataat actgttgatg ggtgtctggt cagagacatc aagaaataac gccggaacat
     3781 tagtgcaggc agcttccaca gcaatggcat cctggtcatc cagcggatag ttaatgatca
     3841 gcccactgac gcgttgcgcg agaagattgt gcaccgccgc tttacaggct tcgacgccgc
     3901 ttcgttctac catcgacacc accacgctgg cacccagttg atcggcgcga gatttaatcg
     3961 ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca
     4021 acgactgttt gcccgccagt tgttgtgcca cgcggttggg aatgtaattc agctccgcca
     4081 tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc
     4141 gggaaacggt ctgataagag acaccggcat actctgcgac atcgtataac gttactggtt
     4201 tcacattcac caccctgaat tgactctctt ccgggcgcta tcatgccata ccgcgaaagg
     4261 ttttgcgcca ttcgatggtg tccgggatct cgacgctc
//
