LOCUS       sequence-63932-e        4380 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 4380)
  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 4380)
  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: pHYDuet216A
FEATURES             Location/Qualifiers
     source          1..4380
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             14..31
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             41..58
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             848..892
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(930..948)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      944..991
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1221..2036)
                     /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(1221..2036)
                     /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     1944..1963
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(2037..2128)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2144..2893)
                     /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(2154..2171)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(2394..2413)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3089..4171)
                     /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     4133..4152
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4172..4249)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     4295..4314
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4295..4313
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    4314..4338
                     /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)."
ORIGIN
        1 catgggcagc agccatcatc atcatcatca cagcagcggc ctggtgccgc gcggcagcct
       61 aggacagtac aaacttatcc tgaacggtaa aaccctgaaa ggtgaaacca ccaccgaagc
      121 tgttgacgct gctaccgcgg aaaaagtttt caaacagtac gctaacgaca acggtgttga
      181 cggtgaatgg acctacgacg acgctaccaa aacctacacc gtaacggaag gatcctgttt
      241 aagctatgaa acggaaatat tgacagtaga atatggatta ttaccgattg gtaaaattgt
      301 agaaaagcgc atcgaatgta ctgtttatag cgttgataat aatggaaata tttatacaca
      361 acctgtagca caatggcacg atcgcggaga acaagaggtg tttgagtatt gtttggaaga
      421 tggttcattg attcgggcaa caaaagacca taagtttatg gtcgacggtc aaatgttgcc
      481 aattgatgaa atatttgaac gtgaattgga tttgatgcgg aaccccggga tcaaaatagc
      541 cacacgtaaa tatttaggca aacaaaatgt ctatgacatt ggagttgagc gcgaccataa
      601 ttttgcactc aaaaatggct tcatagcttc taattgtttc aatggtaccc agtacaaact
      661 tatcctgaac ggtaaaaccc tgaaaggtga aaccaccacc gaagctgttg acgctgctac
      721 cgcggaaaaa gttttcaaac agtacgctaa cgacaacggt gttgacggtg aatggaccta
      781 cgacgacgct accaaaacct acaccgtaac ggaataagct tggctgtttt ggcggctcga
      841 gtctggtaaa gaaaccgctg ctgcgaaatt tgaacgccag cacatggact cgtctactag
      901 cgcagcttaa ttaacctagg ctgctgccac cgctgagcaa taactagcat aaccccttgg
      961 ggcctctaaa cgggtcttga ggggtttttt gctgaaacct caggcatttg agaagcacac
     1021 ggtcacactg cttccggtag tcaataaacc ggtaaaccag caatagacat aagcggctat
     1081 ttaacgaccc tgccctgaac cgacgacaag ctgacgaccg ggtctccgca agtggcactt
     1141 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     1201 atccgctcat gaattaattc ttagaaaaac tcatcgagca tcaaatgaaa ctgcaattta
     1261 ttcatatcag gattatcaat accatatttt tgaaaaagcc gtttctgtaa tgaaggagaa
     1321 aactcaccga ggcagttcca taggatggca agatcctggt atcggtctgc gattccgact
     1381 cgtccaacat caatacaacc tattaatttc ccctcgtcaa aaataaggtt atcaagtgag
     1441 aaatcaccat gagtgacgac tgaatccggt gagaatggca aaagtttatg catttctttc
     1501 cagacttgtt caacaggcca gccattacgc tcgtcatcaa aatcactcgc atcaaccaaa
     1561 ccgttattca ttcgtgattg cgcctgagcg agacgaaata cgcggtcgct gttaaaagga
     1621 caattacaaa caggaatcga atgcaaccgg cgcaggaaca ctgccagcgc atcaacaata
     1681 ttttcacctg aatcaggata ttcttctaat acctggaatg ctgttttccc ggggatcgca
     1741 gtggtgagta accatgcatc atcaggagta cggataaaat gcttgatggt cggaagaggc
     1801 ataaattccg tcagccagtt tagtctgacc atctcatctg taacatcatt ggcaacgcta
     1861 cctttgccat gtttcagaaa caactctggc gcatcgggct tcccatacaa tcgatagatt
     1921 gtcgcacctg attgcccgac attatcgcga gcccatttat acccatataa atcagcatcc
     1981 atgttggaat ttaatcgcgg cctagagcaa gacgtttccc gttgaatatg gctcatactc
     2041 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     2101 tttgaatgta tttagaaaaa taaacaaata ggcatgcagc gctcttccgc ttcctcgctc
     2161 actgactcgc tacgctcggt cgttcgactg cggcgagcgg tgtcagctca ctcaaaagcg
     2221 gtaatacggt tatccacaga atcaggggat aaagccggaa agaacatgtg agcaaaaagc
     2281 aaagcaccgg aagaagccaa cgccgcaggc gtttttccat aggctccgcc cccctgacga
     2341 gcatcacaaa aatcgacgct caagccagag gtggcgaaac ccgacaggac tataaagata
     2401 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     2461 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     2521 ttggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     2581 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     2641 acacgactta tcgccactgg cagcagccat tggtaactga tttagaggac tttgtcttga
     2701 agttatgcac ctgttaaggc taaactgaaa gaacagattt tggtgagtgc ggtcctccaa
     2761 cccacttacc ttggttcaaa gagttggtag ctcagcgaac cttgagaaaa ccaccgttgg
     2821 tagcggtggt ttttctttat ttatgagatg atgaatcaat cggtctatca agtcaacgaa
     2881 cagctattcc gttactctag atttcagtgc aatttatctc ttcaaatgta gcacctgaag
     2941 tcagccccat acgatataag ttgtaattct catgttagtc atgccccgcg cccaccggaa
     3001 ggagctgact gggttgaagg ctctcaaggg catcggtcga gatcccggtg cctaatgagt
     3061 gagctaactt acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     3121 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     3181 ccagggtggt ttttcttttc accagtgaga cgggcaacag ctgattgccc ttcaccgcct
     3241 ggccctgaga gagttgcagc aagcggtcca cgctggtttg ccccagcagg cgaaaatcct
     3301 gtttgatggt ggttaacggc gggatataac atgagctgtc ttcggtatcg tcgtatccca
     3361 ctaccgagat gtccgcacca acgcgcagcc cggactcggt aatggcgcgc attgcgccca
     3421 gcgccatctg atcgttggca accagcatcg cagtgggaac gatgccctca ttcagcattt
     3481 gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc gctatcggct
     3541 gaatttgatt gcgagtgaga tatttatgcc agccagccag acgcagacgc gccgagacag
     3601 aacttaatgg gcccgctaac agcgcgattt gctggtgacc caatgcgacc agatgctcca
     3661 cgcccagtcg cgtaccgtct tcatgggaga aaataatact gttgatgggt gtctggtcag
     3721 agacatcaag aaataacgcc ggaacattag tgcaggcagc ttccacagca atggcatcct
     3781 ggtcatccag cggatagtta atgatcagcc cactgacgcg ttgcgcgaga agattgtgca
     3841 ccgccgcttt acaggcttcg acgccgcttc gttctaccat cgacaccacc acgctggcac
     3901 ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca
     3961 gactggaggt ggcaacgcca atcagcaacg actgtttgcc cgccagttgt tgtgccacgc
     4021 ggttgggaat gtaattcagc tccgccatcg ccgcttccac tttttcccgc gttttcgcag
     4081 aaacgtggct ggcctggttc accacgcggg aaacggtctg ataagagaca ccggcatact
     4141 ctgcgacatc gtataacgtt actggtttca cattcaccac cctgaattga ctctcttccg
     4201 ggcgctatca tgccataccg cgaaaggttt tgcgccattc gatggtgtcc gggatctcga
     4261 cgctctccct tatgcgactc ctgcattagg aaattaatac gactcactat aggggaattg
     4321 tgagcggata acaattcccc tgtagaaata attttgttta actttaataa ggagatatac
//
