LOCUS       sequence-65385-e        5268 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 5268)
  AUTHORS   Aranko AS, Oeemig JS, Kajander T, Iwai H
  TITLE     Intermolecular domain swapping induces intein-mediated protein 
            alternative splicing.
  JOURNAL   Nat Chem Biol. 2013 Aug 25. doi: 10.1038/nchembio.1320.
  PUBMED    23974115
REFERENCE   2  (bases 1 to 5268)
  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: "Nat Chem 
            Biol. 2013 Aug 25. doi: 10.1038/nchembio.1320."
COMMENT     Sequence Label: pSADuet502
FEATURES             Location/Qualifiers
     source          1..5268
                     /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             1822..1866
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1904..1922)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1918..1965
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(2195..3010)
                     /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(2195..3010)
                     /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     2918..2937
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(3011..3102)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3118..3867)
                     /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(3128..3145)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(3368..3387)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4063..5145)
                     /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     5107..5126
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5146..5223)
                     /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 cggttgttta gcaggcgata gtctggtgac attagtagat
      361 agcggactgc aagtgccaat taaggaatta gtaggtaaat ctggttttgc agtttgggca
      421 ttgaacgaag ctacaatgca gctagaaaag gcaattgtta gcaatgcttt ttcgactggt
      481 ataaagcctc tgtttacttt gacaactcga ttggggcgga aaatccgtgc cacaggtaat
      541 cacaagtttt taacaattaa tggctggaaa agacttgatg aattaactcc taaagaacat
      601 ctctgtttac caagaaatct ccccagttct ggtaaacaaa ctatgactta tgccgaagtt
      661 gcattattag ggcatttaat tggcgacggt tgtacattgc cacgtcatgc catacagtac
      721 actaccagag aaatagattt agctcagaat gttgctttct tggcaacaga agtttttgga
      781 gactcaattg ttcctagaat ttcacctgaa cgtgagtggt atcaagttta cttatccgca
      841 gcacagcatc taactcatag tgttagaaat ccaatagcca aatggctaga ctctctgaat
      901 gtttttggtt taaggtctta cgaaaaattt gtacctcgag aattgttctc acaacccaaa
      961 gagttaatag cgtgctttct aagacacctt tggagtacag atggttgtat aaacttaatt
     1021 gcaggaaaaa agccaagacc tattgcattt tatgcaagca gtagtgagag actagctttt
     1081 gatgtacaga cacttttatt aaggcttggt attaatgcaa cactaaggac agttcctcaa
     1141 gttggcaaag gtagaaacca atatcatgta ataattactg gtaagcccga tcttcagtta
     1201 tttattgttc atgttggagc agtaggacaa tataagctac gttcactcca agatattttc
     1261 caacaccttg aaaacagtat tcacaaccct aatagagata taattccgaa ggatatttgg
     1321 aaaatggagg ttgtacctgc gatgcaagct attggcttca ccacacggat attacaagcc
     1381 tccattggag tttcgtattg tggctctact ctttacaaag taaatttaag tcgagaaaga
     1441 gctttaaaag ttggcaacat tgttcaatca tctaaactgg ttactcttgc caaaagtgat
     1501 gtttattggg atgaaatagt ttcaattgaa tatagtggtg aggaagaagt gtttgacctc
     1561 accgttcctg gtttgcataa ttttgttgca aataatatta ttgttcacaa ttccattggt
     1621 acccagtaca aacttatcct gaacggtaaa accctgaaag gtgaaaccac caccgaagct
     1681 gttgacgctg ctaccgcgga aaaagttttc aaacagtacg ctaacgacaa cggtgttgac
     1741 ggtgaatgga cctacgacga cgctaccaaa acctacaccg taacggaata agcttggctg
     1801 ttttggcggc tcgagtctgg taaagaaacc gctgctgcga aatttgaacg ccagcacatg
     1861 gactcgtcta ctagcgcagc ttaattaacc taggctgctg ccaccgctga gcaataacta
     1921 gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa acctcaggca
     1981 tttgagaagc acacggtcac actgcttccg gtagtcaata aaccggtaaa ccagcaatag
     2041 acataagcgg ctatttaacg accctgccct gaaccgacga caagctgacg accgggtctc
     2101 cgcaagtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     2161 acattcaaat atgtatccgc tcatgaatta attcttagaa aaactcatcg agcatcaaat
     2221 gaaactgcaa tttattcata tcaggattat caataccata tttttgaaaa agccgtttct
     2281 gtaatgaagg agaaaactca ccgaggcagt tccataggat ggcaagatcc tggtatcggt
     2341 ctgcgattcc gactcgtcca acatcaatac aacctattaa tttcccctcg tcaaaaataa
     2401 ggttatcaag tgagaaatca ccatgagtga cgactgaatc cggtgagaat ggcaaaagtt
     2461 tatgcatttc tttccagact tgttcaacag gccagccatt acgctcgtca tcaaaatcac
     2521 tcgcatcaac caaaccgtta ttcattcgtg attgcgcctg agcgagacga aatacgcggt
     2581 cgctgttaaa aggacaatta caaacaggaa tcgaatgcaa ccggcgcagg aacactgcca
     2641 gcgcatcaac aatattttca cctgaatcag gatattcttc taatacctgg aatgctgttt
     2701 tcccggggat cgcagtggtg agtaaccatg catcatcagg agtacggata aaatgcttga
     2761 tggtcggaag aggcataaat tccgtcagcc agtttagtct gaccatctca tctgtaacat
     2821 cattggcaac gctacctttg ccatgtttca gaaacaactc tggcgcatcg ggcttcccat
     2881 acaatcgata gattgtcgca cctgattgcc cgacattatc gcgagcccat ttatacccat
     2941 ataaatcagc atccatgttg gaatttaatc gcggcctaga gcaagacgtt tcccgttgaa
     3001 tatggctcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     3061 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggcatg cagcgctctt
     3121 ccgcttcctc gctcactgac tcgctacgct cggtcgttcg actgcggcga gcggtgtcag
     3181 ctcactcaaa agcggtaata cggttatcca cagaatcagg ggataaagcc ggaaagaaca
     3241 tgtgagcaaa aagcaaagca ccggaagaag ccaacgccgc aggcgttttt ccataggctc
     3301 cgcccccctg acgagcatca caaaaatcga cgctcaagcc agaggtggcg aaacccgaca
     3361 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     3421 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     3481 catagctcac gctgttggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     3541 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     3601 tccaacccgg taagacacga cttatcgcca ctggcagcag ccattggtaa ctgatttaga
     3661 ggactttgtc ttgaagttat gcacctgtta aggctaaact gaaagaacag attttggtga
     3721 gtgcggtcct ccaacccact taccttggtt caaagagttg gtagctcagc gaaccttgag
     3781 aaaaccaccg ttggtagcgg tggtttttct ttatttatga gatgatgaat caatcggtct
     3841 atcaagtcaa cgaacagcta ttccgttact ctagatttca gtgcaattta tctcttcaaa
     3901 tgtagcacct gaagtcagcc ccatacgata taagttgtaa ttctcatgtt agtcatgccc
     3961 cgcgcccacc ggaaggagct gactgggttg aaggctctca agggcatcgg tcgagatccc
     4021 ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc cgctttccag
     4081 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     4141 ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca acagctgatt
     4201 gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg tttgccccag
     4261 caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc tgtcttcggt
     4321 atcgtcgtat cccactaccg agatgtccgc accaacgcgc agcccggact cggtaatggc
     4381 gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg gaacgatgcc
     4441 ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt cgccttcccg
     4501 ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag ccagacgcag
     4561 acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt gacccaatgc
     4621 gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa tactgttgat
     4681 gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg cagcttccac
     4741 agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga cgcgttgcgc
     4801 gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta ccatcgacac
     4861 caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa tttgcgacgg
     4921 cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt tgcccgccag
     4981 ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt ccactttttc
     5041 ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg tctgataaga
     5101 gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca ccaccctgaa
     5161 ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcgcc attcgatggt
     5221 gtccgggatc tcgacgctct cccttatgcg actcctgcat taggaaat
//
