LOCUS       sequence-123043-        4540 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  His-Smt3-NpuDnaEN102-CBD.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4540)
  AUTHORS   Guerrero F, Ciragan A, Iwai H
  TITLE     Tandem SUMO fusion vectors for improving soluble protein expression 
            and purification.
  JOURNAL   Protein Expr Purif. 2015 Aug 20. pii: S1046-5928(15)30039-5. doi: 
            10.1016/j.pep.2015.08.019.
  PUBMED    26297996
REFERENCE   2  (bases 1 to 4540)
  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: "Protein 
            Expr Purif. 2015 Aug 20. pii: S1046-5928(15)30039-5. doi: 
            10.1016/j.pep.2015.08.019."
COMMENT     Sequence Label: pHYRSF53
FEATURES             Location/Qualifiers
     source          1..4540
                     /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             124..414
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="SDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPLR
                     RLMEAFAKRQGKEMDSLRFLYDGIRIQADQTPEDLDMEDNDIIEAHREQIGG"
     CDS             910..1065
                     /codon_start=1
                     /gene="Bacillus circulans chiA"
                     /product="chitin binding domain from chitinase A1 (Watanabe
                     et al., 1994)"
                     /label=CBD
                     /translation="TTNPGVSAWQVNTAYTAGQLVTYNGKTYKCLQPHTSLAGWEPSNV
                     PALWQLQ"
     CDS             1094..1138
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1176..1194)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1190..1237
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1467..2282)
                     /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(1467..2282)
                     /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     2190..2209
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(2283..2374)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2390..3139)
                     /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(2400..2417)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(2640..2659)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3335..4417)
                     /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     4379..4398
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4418..4495)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctgta gaaataattt
       61 tgtttaactt taataaggag atataccatg ggcagcagcc atcatcatca tcatcacagc
      121 actagtgact cagaagtcaa tcaagaagct aagccagagg tcaagccaga agtcaagcct
      181 gagactcaca tcaatttaaa ggtgtccgat ggatcttcag aaatcttctt taagatcaaa
      241 aagaccactc ctttaagaag gctgatggaa gcgttcgcta aaagacaggg taaggaaatg
      301 gactccttaa gattcttgta cgacggtatt agaattcaag ctgatcagac ccctgaagat
      361 ttggacatgg aggataacga tattattgag gctcacagag aacagattgg tggatcccat
      421 atgcagtaca aacttatcct gaacggtaaa accctgaaag gtgaaaccac caccgaagct
      481 gttgacgctg ctaccgcgga aaaagttttc aaacagtacg ctaacgacaa cggtgttgac
      541 ggtgaatgga cctacgacga cgctaccaaa acctacaccg taacggaagg atcctgttta
      601 agctatgaaa cggaaatatt gacagtagaa tatggattat taccgattgg taaaattgta
      661 gaaaagcgca tcgaatgtac tgtttatagc gttgataata atggaaatat ttatacacaa
      721 cctgtagcac aatggcacga tcgcggagaa caagaggtgt ttgagtattg tttggaagat
      781 ggttcattga ttcgggcaac aaaagaccat aagtttatga ctgttgatgg tcaaatgttg
      841 ccaattgatg aaatatttga acgtgaattg gatttgatgc gggttgataa tttgccgaac
      901 gaattcggta ccacaaatcc tggtgtatcc gcttggcagg tcaacacagc ttatactgcg
      961 ggacaattgg tcacatataa cggcaagacg tataaatgtt tgcagcccca cacctccttg
     1021 gcaggatggg aaccatccaa cgttcctgcc ttgtggcagc ttcaataagc ttgcggccgc
     1081 actcgagtct ggtaaagaaa ccgctgctgc gaaatttgaa cgccagcaca tggactcgtc
     1141 tactagcgca gcttaattaa cctaggctgc tgccaccgct gagcaataac tagcataacc
     1201 ccttggggcc tctaaacggg tcttgagggg ttttttgctg aaacctcagg catttgagaa
     1261 gcacacggtc acactgcttc cggtagtcaa taaaccggta aaccagcaat agacataagc
     1321 ggctatttaa cgaccctgcc ctgaaccgac gacaagctga cgaccgggtc tccgcaagtg
     1381 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     1441 atatgtatcc gctcatgaat taattcttag aaaaactcat cgagcatcaa atgaaactgc
     1501 aatttattca tatcaggatt atcaatacca tatttttgaa aaagccgttt ctgtaatgaa
     1561 ggagaaaact caccgaggca gttccatagg atggcaagat cctggtatcg gtctgcgatt
     1621 ccgactcgtc caacatcaat acaacctatt aatttcccct cgtcaaaaat aaggttatca
     1681 agtgagaaat caccatgagt gacgactgaa tccggtgaga atggcaaaag tttatgcatt
     1741 tctttccaga cttgttcaac aggccagcca ttacgctcgt catcaaaatc actcgcatca
     1801 accaaaccgt tattcattcg tgattgcgcc tgagcgagac gaaatacgcg gtcgctgtta
     1861 aaaggacaat tacaaacagg aatcgaatgc aaccggcgca ggaacactgc cagcgcatca
     1921 acaatatttt cacctgaatc aggatattct tctaatacct ggaatgctgt tttcccgggg
     1981 atcgcagtgg tgagtaacca tgcatcatca ggagtacgga taaaatgctt gatggtcgga
     2041 agaggcataa attccgtcag ccagtttagt ctgaccatct catctgtaac atcattggca
     2101 acgctacctt tgccatgttt cagaaacaac tctggcgcat cgggcttccc atacaatcga
     2161 tagattgtcg cacctgattg cccgacatta tcgcgagccc atttataccc atataaatca
     2221 gcatccatgt tggaatttaa tcgcggccta gagcaagacg tttcccgttg aatatggctc
     2281 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     2341 tacatatttg aatgtattta gaaaaataaa caaataggca tgcagcgctc ttccgcttcc
     2401 tcgctcactg actcgctacg ctcggtcgtt cgactgcggc gagcggtgtc agctcactca
     2461 aaagcggtaa tacggttatc cacagaatca ggggataaag ccggaaagaa catgtgagca
     2521 aaaagcaaag caccggaaga agccaacgcc gcaggcgttt ttccataggc tccgcccccc
     2581 tgacgagcat cacaaaaatc gacgctcaag ccagaggtgg cgaaacccga caggactata
     2641 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     2701 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     2761 acgctgttgg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     2821 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     2881 ggtaagacac gacttatcgc cactggcagc agccattggt aactgattta gaggactttg
     2941 tcttgaagtt atgcacctgt taaggctaaa ctgaaagaac agattttggt gagtgcggtc
     3001 ctccaaccca cttaccttgg ttcaaagagt tggtagctca gcgaaccttg agaaaaccac
     3061 cgttggtagc ggtggttttt ctttatttat gagatgatga atcaatcggt ctatcaagtc
     3121 aacgaacagc tattccgtta ctctagattt cagtgcaatt tatctcttca aatgtagcac
     3181 ctgaagtcag ccccatacga tataagttgt aattctcatg ttagtcatgc cccgcgccca
     3241 ccggaaggag ctgactgggt tgaaggctct caagggcatc ggtcgagatc ccggtgccta
     3301 atgagtgagc taacttacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     3361 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     3421 tgggcgccag ggtggttttt cttttcacca gtgagacggg caacagctga ttgcccttca
     3481 ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa
     3541 aatcctgttt gatggtggtt aacggcggga tataacatga gctgtcttcg gtatcgtcgt
     3601 atcccactac cgagatgtcc gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg
     3661 cgcccagcgc catctgatcg ttggcaacca gcatcgcagt gggaacgatg ccctcattca
     3721 gcatttgcat ggtttgttga aaaccggaca tggcactcca gtcgccttcc cgttccgcta
     3781 tcggctgaat ttgattgcga gtgagatatt tatgccagcc agccagacgc agacgcgccg
     3841 agacagaact taatgggccc gctaacagcg cgatttgctg gtgacccaat gcgaccagat
     3901 gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat aatactgttg atgggtgtct
     3961 ggtcagagac atcaagaaat aacgccggaa cattagtgca ggcagcttcc acagcaatgg
     4021 catcctggtc atccagcgga tagttaatga tcagcccact gacgcgttgc gcgagaagat
     4081 tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc taccatcgac accaccacgc
     4141 tggcacccag ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca
     4201 gggccagact ggaggtggca acgccaatca gcaacgactg tttgcccgcc agttgttgtg
     4261 ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc ttccactttt tcccgcgttt
     4321 tcgcagaaac gtggctggcc tggttcacca cgcgggaaac ggtctgataa gagacaccgg
     4381 catactctgc gacatcgtat aacgttactg gtttcacatt caccaccctg aattgactct
     4441 cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg gtgtccggga
     4501 tctcgacgct ctcccttatg cgactcctgc attaggaaat
//
