LOCUS       sequence-212887-        5546 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of proISG15 C-term. domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5546)
  AUTHORS   Swatek KN, Aumayr M, Pruneda JN, Visser LJ, Berryman S, Kueck AF, 
            Geurink PP, Ovaa H, van Kuppeveld FJM, Tuthill TJ, Skern T, Komander
            D
  TITLE     Irreversible inactivation of ISG15 by a viral leader protease 
            enables alternative infection detection strategies.
  JOURNAL   Proc Natl Acad Sci U S A. 2018 Mar 6;115(10):2371-2376. doi: 
            10.1073/pnas.1710617115. Epub 2018 Feb 20.
  PUBMED    29463763
REFERENCE   2  (bases 1 to 5546)
  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"; doi: 
            "10.1073/pnas.1710617115"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2018-03-6- 6"; volume: "115"; issue: "10"; pages: "2371-2376"
COMMENT     Sequence Label: mature ISG15 C-term. domain (79-157)
FEATURES             Location/Qualifiers
     source          1..5546
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     106..125
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     209..228
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        209..227
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    228..252
                     /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)."
     RBS             267..289
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             308..325
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             335..358
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             619..636
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(689..707)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      703..750
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      787..1242
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(874..893)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1084..1105
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1334..2149)
                     /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(1334..2149)
                     /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     2057..2076
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2141..2160
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2271..2859
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2760..2779
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3013..3030
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3045..3184
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3110..3129)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3270..3292
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3286..3477)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4286..5368)
                     /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     5330..5349
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5369..5446)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 caaggagatg gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa
       61 acaagcgctc atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat
      121 ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta
      181 gaggatcgag atctcgatcc cgcgaaatta atacgactca ctatagggga attgtgagcg
      241 gataacaatt cccctctaga aataattttg tttaacttta agaaggagat ataccatggg
      301 cagcagccat catcatcatc atcacagcag cggtctggaa gttctgtttc agggcccgga
      361 cgaacctctg agcatcctgg tgaggaataa caagggccgc agcagcacct acgaggtgcg
      421 gctgacgcag accgtggccc acctgaagca gcaagtgagc gggctggagg gtgtgcagga
      481 cgacctgttc tggctgacct tcgaggggaa gcccctggag gaccagctcc cgctggggga
      541 gtacggcctc aagcccctga gcaccgtgtt catgaatctg cgcctgcggg gaggctaaag
      601 ctttctagac catttaaaca ccaccaccac caccactgag atccggctgc taacaaagcc
      661 cgaaaggaag ctgagttggc tgctgccacc gctgagcaat aactagcata accccttggg
      721 gcctctaaac gggtcttgag gggttttttg ctgaaaggag gaactatatc cggattggcg
      781 aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
      841 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
      901 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
      961 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     1021 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     1081 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     1141 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     1201 aatttaacgc gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg
     1261 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     1321 catgaattaa ttcttagaaa aactcatcga gcatcaaatg aaactgcaat ttattcatat
     1381 caggattatc aataccatat ttttgaaaaa gccgtttctg taatgaagga gaaaactcac
     1441 cgaggcagtt ccataggatg gcaagatcct ggtatcggtc tgcgattccg actcgtccaa
     1501 catcaataca acctattaat ttcccctcgt caaaaataag gttatcaagt gagaaatcac
     1561 catgagtgac gactgaatcc ggtgagaatg gcaaaagttt atgcatttct ttccagactt
     1621 gttcaacagg ccagccatta cgctcgtcat caaaatcact cgcatcaacc aaaccgttat
     1681 tcattcgtga ttgcgcctga gcgagacgaa atacgcgatc gctgttaaaa ggacaattac
     1741 aaacaggaat cgaatgcaac cggcgcagga acactgccag cgcatcaaca atattttcac
     1801 ctgaatcagg atattcttct aatacctgga atgctgtttt cccggggatc gcagtggtga
     1861 gtaaccatgc atcatcagga gtacggataa aatgcttgat ggtcggaaga ggcataaatt
     1921 ccgtcagcca gtttagtctg accatctcat ctgtaacatc attggcaacg ctacctttgc
     1981 catgtttcag aaacaactct ggcgcatcgg gcttcccata caatcgatag attgtcgcac
     2041 ctgattgccc gacattatcg cgagcccatt tatacccata taaatcagca tccatgttgg
     2101 aatttaatcg cggcctagag caagacgttt cccgttgaat atggctcata acaccccttg
     2161 tattactgtt tatgtaagca gacagtttta ttgttcatga ccaaaatccc ttaacgtgag
     2221 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     2281 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     2341 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     2401 cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct
     2461 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     2521 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     2581 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     2641 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     2701 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     2761 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     2821 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     2881 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     2941 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     3001 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt
     3061 ctccttacgc atctgtgcgg tatttcacac cgcaatggtg cactctcagt acaatctgct
     3121 ctgatgccgc atagttaagc cagtatacac tccgctatcg ctacgtgact gggtcatggc
     3181 tgcgccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc
     3241 atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc
     3301 gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca tcagcgtggt cgtgaagcga
     3361 ttcacagatg tctgcctgtt catccgcgtc cagctcgttg agtttctcca gaagcgttaa
     3421 tgtctggctt ctgataaagc gggccatgtt aagggcggtt ttttcctgtt tggtcactga
     3481 tgcctccgtg taagggggat ttctgttcat gggggtaatg ataccgatga aacgagagag
     3541 gatgctcacg atacgggtta ctgatgatga acatgcccgg ttactggaac gttgtgaggg
     3601 taaacaactg gcggtatgga tgcggcggga ccagagaaaa atcactcagg gtcaatgcca
     3661 gcgcttcgtt aatacagatg taggtgttcc acagggtagc cagcagcatc ctgcgatgca
     3721 gatccggaac ataatggtgc agggcgctga cttccgcgtt tccagacttt acgaaacacg
     3781 gaaaccgaag accattcatg ttgttgctca ggtcgcagac gttttgcagc agcagtcgct
     3841 tcacgttcgc tcgcgtatcg gtgattcatt ctgctaacca gtaaggcaac cccgccagcc
     3901 tagccgggtc ctcaacgaca ggagcacgat catgcgcacc cgtggggccg ccatgccggc
     3961 gataatggcc tgcttctcgc cgaaacgttt ggtggcggga ccagtgacga aggcttgagc
     4021 gagggcgtgc aagattccga ataccgcaag cgacaggccg atcatcgtcg cgctccagcg
     4081 aaagcggtcc tcgccgaaaa tgacccagag cgctgccggc acctgtccta cgagttgcat
     4141 gataaagaag acagtcataa gtgcggcgac gatagtcatg ccccgcgccc accggaagga
     4201 gctgactggg ttgaaggctc tcaagggcat cggtcgagat cccggtgcct aatgagtgag
     4261 ctaacttaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     4321 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgcca
     4381 gggtggtttt tcttttcacc agtgagacgg gcaacagctg attgcccttc accgcctggc
     4441 cctgagagag ttgcagcaag cggtccacgc tggtttgccc cagcaggcga aaatcctgtt
     4501 tgatggtggt taacggcggg atataacatg agctgtcttc ggtatcgtcg tatcccacta
     4561 ccgagatatc cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt gcgcccagcg
     4621 ccatctgatc gttggcaacc agcatcgcag tgggaacgat gccctcattc agcatttgca
     4681 tggtttgttg aaaaccggac atggcactcc agtcgccttc ccgttccgct atcggctgaa
     4741 tttgattgcg agtgagatat ttatgccagc cagccagacg cagacgcgcc gagacagaac
     4801 ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga tgctccacgc
     4861 ccagtcgcgt accgtcttca tgggagaaaa taatactgtt gatgggtgtc tggtcagaga
     4921 catcaagaaa taacgccgga acattagtgc aggcagcttc cacagcaatg gcatcctggt
     4981 catccagcgg atagttaatg atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg
     5041 ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg ctggcaccca
     5101 gttgatcggc gcgagattta atcgccgcga caatttgcga cggcgcgtgc agggccagac
     5161 tggaggtggc aacgccaatc agcaacgact gtttgcccgc cagttgttgt gccacgcggt
     5221 tgggaatgta attcagctcc gccatcgccg cttccacttt ttcccgcgtt ttcgcagaaa
     5281 cgtggctggc ctggttcacc acgcgggaaa cggtctgata agagacaccg gcatactctg
     5341 cgacatcgta taacgttact ggtttcacat tcaccaccct gaattgactc tcttccgggc
     5401 gctatcatgc cataccgcga aaggttttgc gccattcgat ggtgtccggg atctcgacgc
     5461 tctcccttat gcgactcctg cattaggaag cagcccagta gtaggttgag gccgttgagc
     5521 accgccgccg caaggaatgg tgcatg
//
