LOCUS       sequence-242945-        4398 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Orthogonal NpuDnaB mini-intein intein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4398)
  AUTHORS   Beyer HM, Mikula KM, Li M, Wlodawer A, Iwai H
  TITLE     The crystal structure of the naturally split gp41-1 intein guides 
            the engineering of orthogonal split inteins from cis-splicing 
            inteins.
  JOURNAL   FEBS J. 2019 Oct 30. doi: 10.1111/febs.15113.
  PUBMED    31665813
REFERENCE   2  (bases 1 to 4398)
  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 J. 
            2019 Oct 30. doi: 10.1111/febs.15113."
COMMENT     Sequence Label: pHBDuet140
FEATURES             Location/Qualifiers
     source          1..4398
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(109..1191)
                     /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     1153..1172
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1192..1269)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1315..1334
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1315..1333
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1334..1358
                     /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             1414..1431
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1441..1458
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             2266..2310
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(2348..2366)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2362..2409
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(2639..3454)
                     /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(2639..3454)
                     /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     3362..3381
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(3455..3546)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3562..4311)
                     /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(3572..3589)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(3812..3831)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atgccccgcg cccaccggaa ggagctgact gggttgaagg ctctcaaggg catcggtcga
       61 gatcccggtg cctaatgagt gagctaactt acattaattg cgttgcgctc actgcccgct
      121 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
      181 ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga cgggcaacag
      241 ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca cgctggtttg
      301 ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac atgagctgtc
      361 ttcggtatcg tcgtatccca ctaccgagat gtccgcacca acgcgcagcc cggactcggt
      421 aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg cagtgggaac
      481 gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc
      541 ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc agccagccag
      601 acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt gctggtgacc
      661 caatgcgacc agatgctcca cgcccagtcg cgtaccgtct tcatgggaga aaataatact
      721 gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag tgcaggcagc
      781 ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc cactgacgcg
      841 ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg acgccgcttc gttctaccat
      901 cgacaccacc acgctggcac ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg
      961 cgacggcgcg tgcagggcca gactggaggt ggcaacgcca atcagcaacg actgtttgcc
     1021 cgccagttgt tgtgccacgc ggttgggaat gtaattcagc tccgccatcg ccgcttccac
     1081 tttttcccgc gttttcgcag aaacgtggct ggcctggttc accacgcggg aaacggtctg
     1141 ataagagaca ccggcatact ctgcgacatc gtataacgtt actggtttca cattcaccac
     1201 cctgaattga ctctcttccg ggcgctatca tgccataccg cgaaaggttt tgcgccattc
     1261 gatggtgtcc gggatctcga cgctctccct tatgcgactc ctgcattagg aaattaatac
     1321 gactcactat aggggaattg tgagcggata acaattcccc tgtagaaata attttgttta
     1381 actttaataa ggagatatac catgggcagc agccatcatc atcatcatca cagcagcggc
     1441 ctggtgccgc gcggcagcct aggacagtac aaacttatcc tgaacggtaa aaccctgaaa
     1501 ggtgaaacca ccaccgaagc tgttgacgct gctaccgcgg aaaaagtttt caaacagtac
     1561 gctaacgaca acggtgttga cggtgaatgg acctacgacg acgctaccaa aacctacacc
     1621 gtaacggaag gatccggttg tttagcaggc gatagtctgg tgacattagt agatagcgga
     1681 ctgcaagtgc caattaagga attagtaggt aaatctggtt ttgcagtttg ggcattgaac
     1741 gaagctacaa tgcagctaga aaaggcaatt gttagcaatg ctttttcgac tggtaaaaag
     1801 aagctgtttg aattgacaac tcgattgggg cggaaaatcc gtgccacagg taatcacaag
     1861 tttttaacaa ttaatggctg gaaaagactt gatgaattaa ctcctaaaga acatctcgct
     1921 ttaccccgga attctggtag tgatatttat tgggatgaaa tagtttcaaa ggaatatagt
     1981 ggtaaggaag aagtgtttga cctcaccgtt cctggtttgc ataattttgt tgcaaataat
     2041 attattgttc acaactcggg aggtacccag tacaaactta tcctgaacgg taaaaccctg
     2101 aaaggtgaaa ccaccaccga agctgttgac gctgctaccg cggaaaaagt tttcaaacag
     2161 tacgctaacg acaacggtgt tgacggtgaa tggacctacg acgacgctac caaaacctac
     2221 accgtaacgg aataagcttg gctgttttgg cggctcgagt ctggtaaaga aaccgctgct
     2281 gcgaaatttg aacgccagca catggactcg tctactagcg cagcttaatt aacctaggct
     2341 gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg
     2401 ggttttttgc tgaaacctca ggcatttgag aagcacacgg tcacactgct tccggtagtc
     2461 aataaaccgg taaaccagca atagacataa gcggctattt aacgaccctg ccctgaaccg
     2521 acgacaagct gacgaccggg tctccgcaag tggcactttt cggggaaatg tgcgcggaac
     2581 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga attaattctt
     2641 agaaaaactc atcgagcatc aaatgaaact gcaatttatt catatcagga ttatcaatac
     2701 catatttttg aaaaagccgt ttctgtaatg aaggagaaaa ctcaccgagg cagttccata
     2761 ggatggcaag atcctggtat cggtctgcga ttccgactcg tccaacatca atacaaccta
     2821 ttaatttccc ctcgtcaaaa ataaggttat caagtgagaa atcaccatga gtgacgactg
     2881 aatccggtga gaatggcaaa agtttatgca tttctttcca gacttgttca acaggccagc
     2941 cattacgctc gtcatcaaaa tcactcgcat caaccaaacc gttattcatt cgtgattgcg
     3001 cctgagcgag acgaaatacg cggtcgctgt taaaaggaca attacaaaca ggaatcgaat
     3061 gcaaccggcg caggaacact gccagcgcat caacaatatt ttcacctgaa tcaggatatt
     3121 cttctaatac ctggaatgct gttttcccgg ggatcgcagt ggtgagtaac catgcatcat
     3181 caggagtacg gataaaatgc ttgatggtcg gaagaggcat aaattccgtc agccagttta
     3241 gtctgaccat ctcatctgta acatcattgg caacgctacc tttgccatgt ttcagaaaca
     3301 actctggcgc atcgggcttc ccatacaatc gatagattgt cgcacctgat tgcccgacat
     3361 tatcgcgagc ccatttatac ccatataaat cagcatccat gttggaattt aatcgcggcc
     3421 tagagcaaga cgtttcccgt tgaatatggc tcatactctt cctttttcaa tattattgaa
     3481 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     3541 aacaaatagg catgcagcgc tcttccgctt cctcgctcac tgactcgcta cgctcggtcg
     3601 ttcgactgcg gcgagcggtg tcagctcact caaaagcggt aatacggtta tccacagaat
     3661 caggggataa agccggaaag aacatgtgag caaaaagcaa agcaccggaa gaagccaacg
     3721 ccgcaggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     3781 agccagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     3841 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     3901 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgtt ggtatctcag ttcggtgtag
     3961 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     4021 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     4081 gcagccattg gtaactgatt tagaggactt tgtcttgaag ttatgcacct gttaaggcta
     4141 aactgaaaga acagattttg gtgagtgcgg tcctccaacc cacttacctt ggttcaaaga
     4201 gttggtagct cagcgaacct tgagaaaacc accgttggta gcggtggttt ttctttattt
     4261 atgagatgat gaatcaatcg gtctatcaag tcaacgaaca gctattccgt tactctagat
     4321 ttcagtgcaa tttatctctt caaatgtagc acctgaagtc agccccatac gatataagtt
     4381 gtaattctca tgttagtc
//
