LOCUS       sequence-55939-e        4764 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 4764)
  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 4764)
  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: pSKDuet22
FEATURES             Location/Qualifiers
     source          1..4764
                     /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             1318..1362
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1400..1418)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1414..1461
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1691..2506)
                     /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(1691..2506)
                     /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     2414..2433
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(2507..2598)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2614..3363)
                     /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(2624..2641)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(2864..2883)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3559..4641)
                     /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     4603..4622
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4642..4719)
                     /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 cctctgcgtg gccccggata ccttgataaa taccgacaac
      361 ggaagggtag agatcgggaa gttcgtggag gagtggatga aggaagttgg agagataagc
      421 gaggaaggga tcagctatgc accatgcttc aggaaggtcg agacgttcaa agacgggaag
      481 atagttgaat caccaattag aagggtttgg aaactgagag ctccgaagaa gctcgtgagg
      541 ataaaaacgg agaatggaag gagcatagcc ctaaccaggg agaccaagtt actaacgata
      601 aatgatggag aattatcctg ggttgaggcc ggggaggtta aggtaggaac atacgtggga
      661 acggtaaaaa gtgaaaaaga cgtgattcca ggggctggaa agacaataag ggatgtttca
      721 aaactctaca atatggaaat ggaagttaag gattacttaa ccagggaaga agtgagaaaa
      781 gccattgaaa agcttgaaga aattatgaat ccaatgaata tcaagattcc tggggtgcaa
      841 gaaagttacg aagagctgtt aaggaagctt gaaactacca acgacgagag ggttaggaac
      901 gagacattga tcctcctaag tgacgttagc gatgcccacg agctcgccaa ggaaaaaatc
      961 gagaagatca aagagatcgt gaactcggaa gttcactggg agaaagtcac tgaagtaggg
     1021 gaagtagatg gggttgaata cgtctatgac cttaccgtag agggaagcca caactttgta
     1081 gccaacggat tcatagtcca caacacagct gctggtaccc agtacaaact tatcctgaac
     1141 ggtaaaaccc tgaaaggtga aaccaccacc gaagctgttg acgctgctac cgcggaaaaa
     1201 gttttcaaac agtacgctaa cgacaacggt gttgacggtg aatggaccta cgacgacgct
     1261 accaaaacct acaccgtaac ggaataagct tggctgtttt ggcggctcga gtctggtaaa
     1321 gaaaccgctg ctgcgaaatt tgaacgccag cacatggact cgtctactag cgcagcttaa
     1381 ttaacctagg ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa
     1441 cgggtcttga ggggtttttt gctgaaacct caggcatttg agaagcacac ggtcacactg
     1501 cttccggtag tcaataaacc ggtaaaccag caatagacat aagcggctat ttaacgaccc
     1561 tgccctgaac cgacgacaag ctgacgaccg ggtctccgca agtggcactt ttcggggaaa
     1621 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     1681 gaattaattc ttagaaaaac tcatcgagca tcaaatgaaa ctgcaattta ttcatatcag
     1741 gattatcaat accatatttt tgaaaaagcc gtttctgtaa tgaaggagaa aactcaccga
     1801 ggcagttcca taggatggca agatcctggt atcggtctgc gattccgact cgtccaacat
     1861 caatacaacc tattaatttc ccctcgtcaa aaataaggtt atcaagtgag aaatcaccat
     1921 gagtgacgac tgaatccggt gagaatggca aaagtttatg catttctttc cagacttgtt
     1981 caacaggcca gccattacgc tcgtcatcaa aatcactcgc atcaaccaaa ccgttattca
     2041 ttcgtgattg cgcctgagcg agacgaaata cgcggtcgct gttaaaagga caattacaaa
     2101 caggaatcga atgcaaccgg cgcaggaaca ctgccagcgc atcaacaata ttttcacctg
     2161 aatcaggata ttcttctaat acctggaatg ctgttttccc ggggatcgca gtggtgagta
     2221 accatgcatc atcaggagta cggataaaat gcttgatggt cggaagaggc ataaattccg
     2281 tcagccagtt tagtctgacc atctcatctg taacatcatt ggcaacgcta cctttgccat
     2341 gtttcagaaa caactctggc gcatcgggct tcccatacaa tcgatagatt gtcgcacctg
     2401 attgcccgac attatcgcga gcccatttat acccatataa atcagcatcc atgttggaat
     2461 ttaatcgcgg cctagagcaa gacgtttccc gttgaatatg gctcatactc ttcctttttc
     2521 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     2581 tttagaaaaa taaacaaata ggcatgcagc gctcttccgc ttcctcgctc actgactcgc
     2641 tacgctcggt cgttcgactg cggcgagcgg tgtcagctca ctcaaaagcg gtaatacggt
     2701 tatccacaga atcaggggat aaagccggaa agaacatgtg agcaaaaagc aaagcaccgg
     2761 aagaagccaa cgccgcaggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     2821 aatcgacgct caagccagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     2881 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     2941 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg ttggtatctc
     3001 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     3061 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     3121 tcgccactgg cagcagccat tggtaactga tttagaggac tttgtcttga agttatgcac
     3181 ctgttaaggc taaactgaaa gaacagattt tggtgagtgc ggtcctccaa cccacttacc
     3241 ttggttcaaa gagttggtag ctcagcgaac cttgagaaaa ccaccgttgg tagcggtggt
     3301 ttttctttat ttatgagatg atgaatcaat cggtctatca agtcaacgaa cagctattcc
     3361 gttactctag atttcagtgc aatttatctc ttcaaatgta gcacctgaag tcagccccat
     3421 acgatataag ttgtaattct catgttagtc atgccccgcg cccaccggaa ggagctgact
     3481 gggttgaagg ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt
     3541 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     3601 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt
     3661 ttttcttttc accagtgaga cgggcaacag ctgattgccc ttcaccgcct ggccctgaga
     3721 gagttgcagc aagcggtcca cgctggtttg ccccagcagg cgaaaatcct gtttgatggt
     3781 ggttaacggc gggatataac atgagctgtc ttcggtatcg tcgtatccca ctaccgagat
     3841 gtccgcacca acgcgcagcc cggactcggt aatggcgcgc attgcgccca gcgccatctg
     3901 atcgttggca accagcatcg cagtgggaac gatgccctca ttcagcattt gcatggtttg
     3961 ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc gctatcggct gaatttgatt
     4021 gcgagtgaga tatttatgcc agccagccag acgcagacgc gccgagacag aacttaatgg
     4081 gcccgctaac agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg
     4141 cgtaccgtct tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag
     4201 aaataacgcc ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatccag
     4261 cggatagtta atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt
     4321 acaggcttcg acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc
     4381 ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt
     4441 ggcaacgcca atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat
     4501 gtaattcagc tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct
     4561 ggcctggttc accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc
     4621 gtataacgtt actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatca
     4681 tgccataccg cgaaaggttt tgcgccattc gatggtgtcc gggatctcga cgctctccct
     4741 tatgcgactc ctgcattagg aaat
//
