LOCUS       sequence-331343-        5145 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of Nsp3 ubiquitin-like domain (1-111).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5145)
  TITLE     Savchenko Lab Plasmids
REFERENCE   2  (bases 1 to 5145)
  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"
COMMENT     Sequence Label: pMCSG53_Nsp3_Ubl1
FEATURES             Location/Qualifiers
     source          1..5145
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     916..935
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(955..1032)
                     /gene="lacI"
                     /label=lacI promoter
     tRNA            1172..1247
                     /gene="ileX"
                     /product="E. coli tRNA-Ile recognizing AUA"
                     /label=ileX
     primer_bind     1396..1415
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     tRNA            1517..1593
                     /gene="argU"
                     /product="E. coli tRNA-Arg recognizing AGA and AGG"
                     /label=argU
     primer_bind     1868..1887
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1868..1886
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1887..1911
                     /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             1926..1948
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1959..1976
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2001..2021
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             2427..2444
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2497..2515)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2511..2558
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(2682..2701)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2868..2971
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2972..3040,3041..3832)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3190..3209)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4003..4591
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4492..4511
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4745..4762
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(4842..4861)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     CDS             complement(join(5017..5145,1..954))
                     /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"
ORIGIN
        1 cagctgattg cccttcaccg cctggccctg agagagttgc agcaagcggt ccacgctggt
       61 ttgccccagc aggcgaaaat cctgtttgat ggtggttaac ggcgggatat aacatgagct
      121 gtcttcggta tcgtcgtatc ccactaccga gatatccgca ccaacgcgca gcccggactc
      181 ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg
      241 aacgatgccc tcattcagca tttgcatggt ttgttgaaaa ccggacatgg cactccagtc
      301 gccttcccgt tccgctatcg gctgaatttg attgcgagtg agatatttat gccagccagc
      361 cagacgcaga cgcgccgaga cagaacttaa tgggcccgct aacagcgcga tttgctggtg
      421 acccaatgcg accagatgct ccacgcccag tcgcgtaccg tcttcatggg agaaaataat
      481 actgttgatg ggtgtctggt cagagacatc aagaaataac gccggaacat tagtgcaggc
      541 agcttccaca gcaatggcat cctggtcatc cagcggatag ttaatgatca gcccactgac
      601 gcgttgcgcg agaagattgt gcaccgccgc tttacaggct tcgacgccgc ttcgttctac
      661 catcgacacc accacgctgg cacccagttg atcggcgcga gatttaatcg ccgcgacaat
      721 ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca acgactgttt
      781 gcccgccagt tgttgtgcca cgcggttggg aatgtaattc agctccgcca tcgccgcttc
      841 cactttttcc cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt
      901 ctgataagag acaccggcat actctgcgac atcgtataac gttactggtt tcacattcac
      961 caccctgaat tgactctctt ccgggcgcta tcatgccata ccgcgaaagg ttttgcgcca
     1021 ttcgatggtg tccgggatct cgacgctctc ccttatgcga ctcctgcatt aggaagcagc
     1081 ccagtagtag gttgaggccg ttgagcaccg ccgccgcaag gaatggtgca tgcggattgc
     1141 gacacggagt tactttataa tccgctacca tggcccctta gctcagtggt tagagcaggc
     1201 gactcataat cgcttggtcg ctggttcaag tccagcaggg gccaccagat atagcaaagg
     1261 ctgacgagaa atcgtcagcc tttttgcatg caaggagatg gcgcccaaca gtcccccggc
     1321 cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga agtggcgagc
     1381 ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac ctgtggcgcc
     1441 ggcgagatca aaagccattg actcagcaag ggttgaccgt ataattcacg cgattacacc
     1501 gcattgcggt atcaacgcgc ccttagctca gttggataga gcaacgacct tctaagtcgt
     1561 gggccgcagg ttcgaatcct gcagggcgcg ccattacaat tcaatcagtt acgccttctt
     1621 tatatcctcc ataattccag agtgggacat atttgggaca ttatcaccaa aaatgtcgtc
     1681 tattttcctc gcatgctctg tcaaatgatt aggcgcaagg tgagcatacc tacgaaccat
     1741 ttctatggac tcccatccgc ccatttcctg aagcactgat aatgggacgc ctgactgaat
     1801 ccaccggtga tgccggccac gatgcgtccg gcgtagagga tcgagatcga tctcgatccc
     1861 gcgaaattaa tacgactcac tataggggaa ttgtgagcgg ataacaattc ccctctagaa
     1921 ataattttgt ttaactttaa gaaggagata tacatatgca ccatcatcat catcattctt
     1981 ctggtgtaga tctgggtacc gagaacctgt acttccaatc caatgccgca ccgacaaaag
     2041 ttacatttgg agacgatacc gtgatcgaag ttcagggcta caaaagcgtg aacatcacct
     2101 tcgagctgga tgaacgtatc gataaagtgc tgaacgagaa atgcagcgca tataccgtgg
     2161 aactgggtac cgaagtgaac gaatttgcct gtgttgttgc agatgcagtg atcaaaacct
     2221 tacagccggt tagcgaactg ctgacacctt taggcattga tctggatgaa tggagcatgg
     2281 caacctatta tctgttcgac gaaagcggcg agttcaaact ggcatcacac atgtattgca
     2341 gcttctatcc gcctgatgaa taacattgga agtggataac ggatccgaat tcgagctccg
     2401 tcgacaagct tgcggccgca ctcgagcacc accaccacca ccactgagat ccggctgcta
     2461 acaaagcccg aaaggaagct gagttggctg ctgccaccgc tgagcaataa ctagcataac
     2521 cccttggggc ctctaaacgg gtcttgaggg gttttttgct gaaaggagga actatatccg
     2581 gattggcgaa tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     2641 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     2701 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     2761 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     2821 ttcacattaa cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     2881 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     2941 aaatgcttca ataatttgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     3001 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     3061 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     3121 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt
     3181 taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg
     3241 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
     3301 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
     3361 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
     3421 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
     3481 cataccaaac gacgagcgtg acaccacgat gcctgcagca atggcaacaa cgttgcgcaa
     3541 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
     3601 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
     3661 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga
     3721 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
     3781 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
     3841 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     3901 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     3961 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     4021 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     4081 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     4141 aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     4201 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     4261 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     4321 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     4381 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     4441 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     4501 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     4561 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     4621 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     4681 ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga
     4741 gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg atgcggtatt ttctccttac
     4801 gcatctgtgc ggtatttcac accgcaatgg tgcactctca gtacaatctg ctctgatgcc
     4861 gcatagttaa gccagtatac actccgctat cgctacgtga ctgtagtcat gccccgcgcc
     4921 caccggaagg agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc
     4981 taatgagtga gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     5041 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     5101 attgggcgcc agggtggttt ttcttttcac cagtgagacg ggcaa
//
