LOCUS       sequence-57155-e        4506 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 4506)
  AUTHORS   Ellila S, Jurvansuu JM, Iwai H
  TITLE     Evaluation and comparison of protein splicing by exogenous inteins 
            with foreign exteins in Escherichia coli.
  JOURNAL   FEBS Lett. 2011 Oct 10.
  PUBMED    22001202
REFERENCE   2  (bases 1 to 4506)
  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. 
            2011 Oct 10."
COMMENT     Sequence Label: pSKDuet27
FEATURES             Location/Qualifiers
     source          1..4506
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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             100..117
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             127..144
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1060..1104
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1142..1160)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1156..1203
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1433..2248)
                     /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(1433..2248)
                     /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     2156..2175
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(2249..2340)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2356..3105)
                     /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(2366..2383)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(2606..2625)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3301..4383)
                     /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     4345..4364
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4384..4461)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctgta gaaataattt
       61 tgtttaactt taataaggag atataccatg ggcagcagcc atcatcatca tcatcacagc
      121 agcggcctgg tgccgcgcgg cagcctagga cagtacaaac ttatcctgaa cggtaaaacc
      181 ctgaaaggtg aaaccaccac cgaagctgtt gacgctgcta ccgcggaaaa agttttcaaa
      241 cagtacgcta acgacaacgg tgttgacggt gaatggacct acgacgacgc taccaaaacc
      301 tacaccgtaa cggaaggatc caaatgtgtg tctggcgata caccggtact tctggatgcc
      361 ggcgagagga ggataggcga cctgttcatg gaggccattc aggaccaaaa gaacgcggtc
      421 gaaataggcc agaacgaaga gatagtccgg ctccatgatc cgctgcgcat atattccatg
      481 gtcggttctg aaatagtcga aagcgtctct cacgccatat atcacggaaa gagcaatgcc
      541 attgtaaccg ttaggacgga gaatggaaga gaggtcaggg tgacacctgt ccacaaactc
      601 tttgttaaaa ttggaaactc tgtaatcgag aggccagcct cagaggtgaa tgagggcgat
      661 gaaatagcat gcgcaagcgt aagtgagaac ggtgattccc aaaccgtcac cacaacgctg
      721 gtattgacat tcgatagagt ggtatcaaag gaaatgcata gcggcgtatt cgatgtctac
      781 gatctgatgg ttccggatta tggatacaac ttcataggcg gaaatggcct catagtcctt
      841 cacaacaccg tgataggtac ccagtacaaa cttatcctga acggtaaaac cctgaaaggt
      901 gaaaccacca ccgaagctgt tgacgctgct accgcggaaa aagttttcaa acagtacgct
      961 aacgacaacg gtgttgacgg tgaatggacc tacgacgacg ctaccaaaac ctacaccgta
     1021 acggaataag cttggctgtt ttggcggctc gagtctggta aagaaaccgc tgctgcgaaa
     1081 tttgaacgcc agcacatgga ctcgtctact agcgcagctt aattaaccta ggctgctgcc
     1141 accgctgagc aataactagc ataacccctt ggggcctcta aacgggtctt gaggggtttt
     1201 ttgctgaaac ctcaggcatt tgagaagcac acggtcacac tgcttccggt agtcaataaa
     1261 ccggtaaacc agcaatagac ataagcggct atttaacgac cctgccctga accgacgaca
     1321 agctgacgac cgggtctccg caagtggcac ttttcgggga aatgtgcgcg gaacccctat
     1381 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgaattaat tcttagaaaa
     1441 actcatcgag catcaaatga aactgcaatt tattcatatc aggattatca ataccatatt
     1501 tttgaaaaag ccgtttctgt aatgaaggag aaaactcacc gaggcagttc cataggatgg
     1561 caagatcctg gtatcggtct gcgattccga ctcgtccaac atcaatacaa cctattaatt
     1621 tcccctcgtc aaaaataagg ttatcaagtg agaaatcacc atgagtgacg actgaatccg
     1681 gtgagaatgg caaaagttta tgcatttctt tccagacttg ttcaacaggc cagccattac
     1741 gctcgtcatc aaaatcactc gcatcaacca aaccgttatt cattcgtgat tgcgcctgag
     1801 cgagacgaaa tacgcggtcg ctgttaaaag gacaattaca aacaggaatc gaatgcaacc
     1861 ggcgcaggaa cactgccagc gcatcaacaa tattttcacc tgaatcagga tattcttcta
     1921 atacctggaa tgctgttttc ccggggatcg cagtggtgag taaccatgca tcatcaggag
     1981 tacggataaa atgcttgatg gtcggaagag gcataaattc cgtcagccag tttagtctga
     2041 ccatctcatc tgtaacatca ttggcaacgc tacctttgcc atgtttcaga aacaactctg
     2101 gcgcatcggg cttcccatac aatcgataga ttgtcgcacc tgattgcccg acattatcgc
     2161 gagcccattt atacccatat aaatcagcat ccatgttgga atttaatcgc ggcctagagc
     2221 aagacgtttc ccgttgaata tggctcatac tcttcctttt tcaatattat tgaagcattt
     2281 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     2341 taggcatgca gcgctcttcc gcttcctcgc tcactgactc gctacgctcg gtcgttcgac
     2401 tgcggcgagc ggtgtcagct cactcaaaag cggtaatacg gttatccaca gaatcagggg
     2461 ataaagccgg aaagaacatg tgagcaaaaa gcaaagcacc ggaagaagcc aacgccgcag
     2521 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagccag
     2581 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     2641 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     2701 ggaagcgtgg cgctttctca tagctcacgc tgttggtatc tcagttcggt gtaggtcgtt
     2761 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     2821 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     2881 attggtaact gatttagagg actttgtctt gaagttatgc acctgttaag gctaaactga
     2941 aagaacagat tttggtgagt gcggtcctcc aacccactta ccttggttca aagagttggt
     3001 agctcagcga accttgagaa aaccaccgtt ggtagcggtg gtttttcttt atttatgaga
     3061 tgatgaatca atcggtctat caagtcaacg aacagctatt ccgttactct agatttcagt
     3121 gcaatttatc tcttcaaatg tagcacctga agtcagcccc atacgatata agttgtaatt
     3181 ctcatgttag tcatgccccg cgcccaccgg aaggagctga ctgggttgaa ggctctcaag
     3241 ggcatcggtc gagatcccgg tgcctaatga gtgagctaac ttacattaat tgcgttgcgc
     3301 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     3361 cgcgcgggga gaggcggttt gcgtattggg cgccagggtg gtttttcttt tcaccagtga
     3421 gacgggcaac agctgattgc ccttcaccgc ctggccctga gagagttgca gcaagcggtc
     3481 cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata
     3541 acatgagctg tcttcggtat cgtcgtatcc cactaccgag atgtccgcac caacgcgcag
     3601 cccggactcg gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg caaccagcat
     3661 cgcagtggga acgatgccct cattcagcat ttgcatggtt tgttgaaaac cggacatggc
     3721 actccagtcg ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga gatatttatg
     3781 ccagccagcc agacgcagac gcgccgagac agaacttaat gggcccgcta acagcgcgat
     3841 ttgctggtga cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt cttcatggga
     3901 gaaaataata ctgttgatgg gtgtctggtc agagacatca agaaataacg ccggaacatt
     3961 agtgcaggca gcttccacag caatggcatc ctggtcatcc agcggatagt taatgatcag
     4021 cccactgacg cgttgcgcga gaagattgtg caccgccgct ttacaggctt cgacgccgct
     4081 tcgttctacc atcgacacca ccacgctggc acccagttga tcggcgcgag atttaatcgc
     4141 cgcgacaatt tgcgacggcg cgtgcagggc cagactggag gtggcaacgc caatcagcaa
     4201 cgactgtttg cccgccagtt gttgtgccac gcggttggga atgtaattca gctccgccat
     4261 cgccgcttcc actttttccc gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg
     4321 ggaaacggtc tgataagaga caccggcata ctctgcgaca tcgtataacg ttactggttt
     4381 cacattcacc accctgaatt gactctcttc cgggcgctat catgccatac cgcgaaaggt
     4441 tttgcgccat tcgatggtgt ccgggatctc gacgctctcc cttatgcgac tcctgcatta
     4501 ggaaat
//
