LOCUS       sequence-55938-e        4488 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 4488)
  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 4488)
  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: pSKDuet21
FEATURES             Location/Qualifiers
     source          1..4488
                     /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             1042..1086
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1124..1142)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1138..1185
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1415..2230)
                     /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(1415..2230)
                     /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     2138..2157
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(2231..2322)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2338..3087)
                     /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(2348..2365)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(2588..2607)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3283..4365)
                     /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     4327..4346
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4366..4443)
                     /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 caagtgcgtt gactacgata cagaagtcct acttggagat
      361 gggagaaaga ggaagatagg agagatcgtt gaggaagcga taaagaaagc agaaaaagag
      421 ggaaaacttg gaagggttga cgatggcttc tacgctccaa taaacctgga gctgtacgct
      481 ttagatgtga gaacccttaa ggtcaggaag gtcaaagctg acatagcgtg gaagagaaca
      541 accccggaga agatgttgag gattagaact aagaggggta gggaaattag ggtaacccca
      601 acgcacccat tcttcacgtt agaggaggga aggataaaga cgaagaaagc ttatgagcta
      661 aaggttgggg agaagatagc cacccccagg gaggaagcac ccgaagctga gatattttgg
      721 gatgaagttg ttgagattga ggagtacaag cccaataact cttgggttta cgatctccaa
      781 gttccagagc accacaactt catagcgaac ggaatattcg tgcacaatag ccaactcggt
      841 acccagtaca aacttatcct gaacggtaaa accctgaaag gtgaaaccac caccgaagct
      901 gttgacgctg ctaccgcgga aaaagttttc aaacagtacg ctaacgacaa cggtgttgac
      961 ggtgaatgga cctacgacga cgctaccaaa acctacaccg taacggaata agcttggctg
     1021 ttttggcggc tcgagtctgg taaagaaacc gctgctgcga aatttgaacg ccagcacatg
     1081 gactcgtcta ctagcgcagc ttaattaacc taggctgctg ccaccgctga gcaataacta
     1141 gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa acctcaggca
     1201 tttgagaagc acacggtcac actgcttccg gtagtcaata aaccggtaaa ccagcaatag
     1261 acataagcgg ctatttaacg accctgccct gaaccgacga caagctgacg accgggtctc
     1321 cgcaagtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     1381 acattcaaat atgtatccgc tcatgaatta attcttagaa aaactcatcg agcatcaaat
     1441 gaaactgcaa tttattcata tcaggattat caataccata tttttgaaaa agccgtttct
     1501 gtaatgaagg agaaaactca ccgaggcagt tccataggat ggcaagatcc tggtatcggt
     1561 ctgcgattcc gactcgtcca acatcaatac aacctattaa tttcccctcg tcaaaaataa
     1621 ggttatcaag tgagaaatca ccatgagtga cgactgaatc cggtgagaat ggcaaaagtt
     1681 tatgcatttc tttccagact tgttcaacag gccagccatt acgctcgtca tcaaaatcac
     1741 tcgcatcaac caaaccgtta ttcattcgtg attgcgcctg agcgagacga aatacgcggt
     1801 cgctgttaaa aggacaatta caaacaggaa tcgaatgcaa ccggcgcagg aacactgcca
     1861 gcgcatcaac aatattttca cctgaatcag gatattcttc taatacctgg aatgctgttt
     1921 tcccggggat cgcagtggtg agtaaccatg catcatcagg agtacggata aaatgcttga
     1981 tggtcggaag aggcataaat tccgtcagcc agtttagtct gaccatctca tctgtaacat
     2041 cattggcaac gctacctttg ccatgtttca gaaacaactc tggcgcatcg ggcttcccat
     2101 acaatcgata gattgtcgca cctgattgcc cgacattatc gcgagcccat ttatacccat
     2161 ataaatcagc atccatgttg gaatttaatc gcggcctaga gcaagacgtt tcccgttgaa
     2221 tatggctcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     2281 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggcatg cagcgctctt
     2341 ccgcttcctc gctcactgac tcgctacgct cggtcgttcg actgcggcga gcggtgtcag
     2401 ctcactcaaa agcggtaata cggttatcca cagaatcagg ggataaagcc ggaaagaaca
     2461 tgtgagcaaa aagcaaagca ccggaagaag ccaacgccgc aggcgttttt ccataggctc
     2521 cgcccccctg acgagcatca caaaaatcga cgctcaagcc agaggtggcg aaacccgaca
     2581 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     2641 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     2701 catagctcac gctgttggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     2761 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     2821 tccaacccgg taagacacga cttatcgcca ctggcagcag ccattggtaa ctgatttaga
     2881 ggactttgtc ttgaagttat gcacctgtta aggctaaact gaaagaacag attttggtga
     2941 gtgcggtcct ccaacccact taccttggtt caaagagttg gtagctcagc gaaccttgag
     3001 aaaaccaccg ttggtagcgg tggtttttct ttatttatga gatgatgaat caatcggtct
     3061 atcaagtcaa cgaacagcta ttccgttact ctagatttca gtgcaattta tctcttcaaa
     3121 tgtagcacct gaagtcagcc ccatacgata taagttgtaa ttctcatgtt agtcatgccc
     3181 cgcgcccacc ggaaggagct gactgggttg aaggctctca agggcatcgg tcgagatccc
     3241 ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc cgctttccag
     3301 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     3361 ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca acagctgatt
     3421 gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg tttgccccag
     3481 caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc tgtcttcggt
     3541 atcgtcgtat cccactaccg agatgtccgc accaacgcgc agcccggact cggtaatggc
     3601 gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg gaacgatgcc
     3661 ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt cgccttcccg
     3721 ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag ccagacgcag
     3781 acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt gacccaatgc
     3841 gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa tactgttgat
     3901 gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg cagcttccac
     3961 agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga cgcgttgcgc
     4021 gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta ccatcgacac
     4081 caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa tttgcgacgg
     4141 cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt tgcccgccag
     4201 ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt ccactttttc
     4261 ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg tctgataaga
     4321 gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca ccaccctgaa
     4381 ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcgcc attcgatggt
     4441 gtccgggatc tcgacgctct cccttatgcg actcctgcat taggaaat
//
