LOCUS       sequence-212819-        6633 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of proISG15.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6633)
  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 6633)
  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: ISG15 C-term. Domain deltaC intein/chitin bind...
FEATURES             Location/Qualifiers
     source          1..6633
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      115..201
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     336..355
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        336..354
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    355..379
                     /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             394..416
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             725..1318
                     /codon_start=1
                     /gene="Mycobacterium xenopi gyrA"
                     /product="modified GyrA intein (Southworth et al., 1999)"
                     /label=Mxe GyrA intein
                     /note="N-terminal cleavage of this intein can be induced by
                     thiols."
                     /translation="CITGDALVALPEGESVRIADIVPGARPNSDNAIDLKVLDRHGNPV
                     LADRLFHSGEHPVYTVRTVEGLRVTGTANHPLLCLVDVAGVPTLLWKLIDEIKPGDYAV
                     IQRSAFSVDCAGFARGKPEFAPTTYTVGVPGLVRFLEAHHRDPDAQAIADELTDGRFYY
                     AKVASVTDAGVQPVYSLRVDTADHAFITNGFVSHA"
     CDS             1349..1504
                     /codon_start=1
                     /gene="Bacillus circulans chiA"
                     /product="chitin binding domain from chitinase A1 (Watanabe
                     et al., 1994)"
                     /label=CBD
                     /translation="TTNPGVSAWQVNTAYTAGQLVTYNGKTYKCLQPHTSLAGWEPSNV
                     PALWQLQ"
     terminator      1583..1630
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        1690..1794
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1795..1863,1864..2655)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2013..2032)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(2697..3210)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2892..2913)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3104..3123
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      3321..3909
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3810..3829
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4063..4080
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(4102..4121)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4262..4284
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4278..4469)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5026..6108)
                     /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     6070..6089
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6109..6186)
                     /gene="lacI"
                     /label=lacI promoter
     terminator      6339..6425
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      6522..6608
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ctcctgagta ggacaaatcc gccgggagcg gatttgaacg ttgcgaagca acggcccgga
       61 gggtggcggg caggacgccc gccataaact gccaggaatt aattccccag gcatcaaata
      121 aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcggtgaac
      181 gctctcctga gtaggacaaa tccgccggga gcggatttga acgttgcgaa gcaacggccc
      241 ggagggtggc gggcaggacg cccgccataa actgccagga attggggatc ggaattaatt
      301 cccggtttaa accggggatc tcgatcccgc gaaattaata cgactcacta taggggaatt
      361 gtgagcggat aacaattccc ctctagaaat aattttgttt aactttaaga aggagatata
      421 catatggcta gctcgcgagt cgacggcggc cgcgaattcc tcgagggctc ttcctgcgga
      481 agagcaacca cctatggatg aaccgctgag cattctggtg cgtaataaca aaggtcgtag
      541 cagcacctat gaagttcgtc tgacccagac agttgcacat ctgaaacagc aggttagcgg
      601 tctggaaggt gttcaggatg acctgttttg gctgaccttt gaaggtaaac cgctggaaga
      661 tcagctgccg ctgggtgaat atggtctgaa accgctgtca accgttttta tgaatctgcg
      721 tctgtgcatc acgggagatg cactagttgc cctacccgag ggcgagtcgg tacgcatcgc
      781 cgacatcgtg ccgggtgcgc ggcccaacag tgacaacgcc atcgacctga aagtccttga
      841 ccggcatggc aatcccgtgc tcgccgaccg gctgttccac tccggcgagc atccggtgta
      901 cacggtgcgt acggtcgaag gtctgcgtgt gacgggcacc gcgaaccacc cgttgttgtg
      961 tttggtcgac gtcgccgggg tgccgaccct gctgtggaag ctgatcgacg aaatcaagcc
     1021 gggcgattac gcggtgattc aacgcagcgc attcagcgtc gactgtgcag gttttgcccg
     1081 cgggaaaccc gaatttgcgc ccacaaccta cacagtcggc gtccctggac tggtgcgttt
     1141 cttggaagca caccaccgag acccggacgc ccaagctatc gccgacgagc tgaccgacgg
     1201 gcggttctac tacgcgaaag tcgccagtgt caccgacgcc ggcgtgcagc cggtgtatag
     1261 ccttcgtgtc gacacggcag accacgcgtt tatcacgaac gggttcgtca gccacgctac
     1321 tggcctcacc ggtctgaact caggcctcac gacaaatcct ggtgtatccg cttggcaggt
     1381 caacacagct tatactgcgg gacaattggt cacatataac ggcaagacgt ataaatgttt
     1441 gcagccccac acctccttgg caggatggga accatccaac gttcctgcct tgtggcagct
     1501 tcaatgactg caggaagggg atccggctgc taacaaagcc cgaaaggaag ctgagttggc
     1561 tgctgccacc gctgagcaat aactagcata accccttggg gcctctaaac gggtcttgag
     1621 gggttttttg ctgaaaggag gaactatatc cggataacta cgtcaggtgg cacttttcgg
     1681 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     1741 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
     1801 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
     1861 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
     1921 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
     1981 cgtttcccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtgtt
     2041 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     2101 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     2161 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     2221 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     2281 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
     2341 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     2401 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     2461 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     2521 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     2581 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     2641 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaccc
     2701 cggttgataa tcagaaaagc cccaaaaaca ggaagattgt ataagcaaat atttaaattg
     2761 taaacgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta
     2821 accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagccc gagatagggt
     2881 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca
     2941 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca cccaaatcaa
     3001 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat
     3061 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag
     3121 gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg
     3181 ccgcgcttaa tgcgccgcta cagggcgcgt aaaaggatct aggtgaagat cctttttgat
     3241 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     3301 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     3361 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     3421 tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag
     3481 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     3541 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     3601 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     3661 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     3721 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     3781 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     3841 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     3901 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     3961 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
     4021 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
     4081 gaagctatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatac
     4141 actccgctat cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc aacacccgct
     4201 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc
     4261 tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg
     4321 cggtaaagct catcagcgtg gtcgtgcagc gattcacaga tgtctgcctg ttcatccgcg
     4381 tccagctcgt tgagtttctc cagaagcgtt aatgtctggc ttctgataaa gcgggccatg
     4441 ttaagggcgg ttttttcctg tttggtcact gatgcctccg tgtaaggggg atttctgttc
     4501 atgggggtaa tgataccgat gaaacgagag aggatgctca cgatacgggt tactgatgat
     4561 gaacatgccc ggttactgga acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg
     4621 gaccagagaa aaatcactca gggtcaatgc cagccgaacg ccagcaagac gtagcccagc
     4681 gcgtcggccg ccatgccggc gataatggcc tgcttctcgc cgaaacgttt ggtggcggga
     4741 ccagtgacga aggcttgagc gagggcgtgc aagattccga ataccgcaag cgacaggccg
     4801 atcatcgtcg cgctccagcg aaagcggtcc tcgccgaaaa tgacccagag cgctgccggc
     4861 acctgtccta cgagttgcat gataaagaag acagtcataa gtgcggcgac gatagtcatg
     4921 ccccgcgccc accggaagga gctgactggg ttgaaggctc tcaagggcat cggtcgagat
     4981 cccggtgcct aatgagtgag ctaacttaca ttaattgcgt tgcgctcact gcccgctttc
     5041 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     5101 ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg
     5161 attgcccttc accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc
     5221 cagcaggcga aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc
     5281 ggtatcgtcg tatcccacta ccgagatatc cgcaccaacg cgcagcccgg actcggtaat
     5341 ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat
     5401 gccctcattc agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc
     5461 ccgttccgct atcggctgaa tttgattgcg agtgagatat ttatgccagc cagccagacg
     5521 cagacgcgcc gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa
     5581 tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt
     5641 gatgggtgtc tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc
     5701 cacagcaatg gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg
     5761 cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga
     5821 caccaccacg ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga
     5881 cggcgcgtgc agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc
     5941 cagttgttgt gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt
     6001 ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata
     6061 agagacaccg gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct
     6121 gaattgactc tcttccgggc gctatcatgc cataccgcga aaggttttgc gccattcgat
     6181 ggtgtccggg atctcgacgc tctcccttat gcgactcctg cattaggaag cagcccagta
     6241 gtaggttgag gccgttgagc accgccgccg caaggaatgg tgcatgccgg catgccgccc
     6301 tttcgtcttc aagaattaat tcccaattcc ccaggcatca aataaaacga aaggctcagt
     6361 cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc ctgagtagga
     6421 caaatccgcc gggagcggat ttgaacgttg cgaagcaacg gcccggaggg tggcgggcag
     6481 gacgcccgcc ataaactgcc aggaattaat tccccaggca tcaaataaaa cgaaaggctc
     6541 agtcgaaaga ctgggccttt cgttttatct gttgtttgtc ggtgaacgct ctcctgagta
     6601 ggacaaatcc gccgggagcg gatttgaacg ttg
//
