LOCUS       sequence-267362-        6199 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses SUMO3-tagged ORF24-NTD-Strep in bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6199)
  AUTHORS   Castaneda AF, Didychuk AL, Louder RK, McCollum CO, Davis ZH, Nogales
            E, Glaunsinger BA
  TITLE     The gammaherpesviral TATA-box-binding protein directly interacts 
            with the CTD of host RNA Pol II to direct late gene transcription.
  JOURNAL   PLoS Pathog. 2020 Sep 4;16(9):e1008843. doi: 
            10.1371/journal.ppat.1008843. eCollection 2020 Sep.
  PUBMED    32886723
REFERENCE   2  (bases 1 to 6199)
  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: "PLoS 
            Pathog."; date: "2020-09-4"; volume: "16(9):e1008843. doi"; pages: "
            10.1371/journal.ppat.1008843. eCollection 2020 Sep"
COMMENT     Sequence Label: p6H-SUMO3-ORF24-NTD-Strep
FEATURES             Location/Qualifiers
     source          1..6199
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     21..40
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        21..39
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    40..64
                     /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             79..101
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             114..131
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             144..413
                     /codon_start=1
                     /product="human small ubiquitin-related modifier 3"
                     /label=SUMO3
                     /translation="EEKPKEGVKTENDHINLKVAGQDGSVVQFKIKRHTPLSKLMKAYC
                     ERQGLSMRQIRFRFDGQPINETDTPAQLEMEDEDTIDVFQQQTGG"
     CDS             1014..1037
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1080..1097
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      1164..1211
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1248..1703
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1335..1354)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1545..1566
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1796..2611)
                     /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(1796..2611)
                     /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     2519..2538
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2603..2622
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2733..3321
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3222..3241
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3475..3492
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3507..3649
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3575..3594)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3735..3757
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3751..3942)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4751..5833)
                     /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     5795..5814
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5834..5911)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6117..6136
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 agatctcgat cccgcgaaat taatacgact cactataggg gaattgtgag cggataacaa
       61 ttcccctcta gaaataattt tgtttaactt taagaaggag atataccatg ggtcatcacc
      121 atcatcatca cgggtccctg caggaggaga agcccaagga gggtgtgaag acagagaatg
      181 accacatcaa cctgaaggtg gccgggcagg acggctccgt ggtgcagttc aagatcaaga
      241 ggcacacgcc gctgagcaag ctgatgaagg cctactgcga gaggcagggc ttgtcaatga
      301 ggcagatcag attcaggttc gacgggcagc caatcaatga aactgacact ccagcacagc
      361 tggagatgga ggacgaggac accatcgacg tgttccagca gcagacggga ggggcagcgc
      421 tcgagggccc cctactactg ccaccgagcg cctccctgac gacgagtccg cagaccacgt
      481 gttatcaagc gacttgggaa tcacagctgg aaatattctg ctgtctggcc accaactcgc
      541 acctgcaggc agagctgacc ttagaaggtc ttgataagat gatgcagccc gagcccacct
      601 ttttcgcctg cagagcgata cgcagactac tcctggggga acgcctccac ccttttatac
      661 atcaagaagg gactcttttg ggaaaagtgg gtcgacggta cagcggcgaa ggtttaataa
      721 ttgacggtgg tggagtgttt acgcgcggac agatagacac cgacaactac ctacctgcgg
      781 tgggatcatg ggaacttacc gatgattgtg ataaaccctg cgaattcagg gagctacgct
      841 cgctgtatct tcccgcgcta ctaacgtgca ccatatgtta caaagccatg ttcaggatag
      901 tgtgcaggta cctggagttc tgggagttcg aacagtgttt tcatgcgttt ctggcggtgt
      961 tgccccatag tctacaaccc acaatctatc aaaattattt tgcactcctg gagtggagcc
     1021 atccgcagtt tgaaaaataa aattcgagct ccgtcgacaa gcttgcggcc gcactcgagc
     1081 accaccacca ccaccactga gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg
     1141 ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa cgggtcttga
     1201 ggggtttttt gctgaaagga ggaactatat ccggattggc gaatgggacg cgccctgtag
     1261 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     1321 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     1381 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     1441 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     1501 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     1561 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     1621 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa
     1681 caaaatatta acgtttacaa tttcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     1741 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgaatt aattcttaga
     1801 aaaactcatc gagcatcaaa tgaaactgca atttattcat atcaggatta tcaataccat
     1861 atttttgaaa aagccgtttc tgtaatgaag gagaaaactc accgaggcag ttccatagga
     1921 tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc aacatcaata caacctatta
     1981 atttcccctc gtcaaaaata aggttatcaa gtgagaaatc accatgagtg acgactgaat
     2041 ccggtgagaa tggcaaaagt ttatgcattt ctttccagac ttgttcaaca ggccagccat
     2101 tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt attcattcgt gattgcgcct
     2161 gagcgagacg aaatacgcga tcgctgttaa aaggacaatt acaaacagga atcgaatgca
     2221 accggcgcag gaacactgcc agcgcatcaa caatattttc acctgaatca ggatattctt
     2281 ctaatacctg gaatgctgtt ttcccgggga tcgcagtggt gagtaaccat gcatcatcag
     2341 gagtacggat aaaatgcttg atggtcggaa gaggcataaa ttccgtcagc cagtttagtc
     2401 tgaccatctc atctgtaaca tcattggcaa cgctaccttt gccatgtttc agaaacaact
     2461 ctggcgcatc gggcttccca tacaatcgat agattgtcgc acctgattgc ccgacattat
     2521 cgcgagccca tttataccca tataaatcag catccatgtt ggaatttaat cgcggcctag
     2581 agcaagacgt ttcccgttga atatggctca taacacccct tgtattactg tttatgtaag
     2641 cagacagttt tattgttcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     2701 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     2761 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     2821 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc
     2881 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     2941 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     3001 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     3061 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     3121 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     3181 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     3241 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     3301 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     3361 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     3421 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag
     3481 tcagtgagcg aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc
     3541 ggtatttcac accgcatata tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     3601 taagccagta tacactccgc tatcgctacg tgactgggtc atggctgcgc cccgacaccc
     3661 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     3721 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     3781 cgcgaggcag ctgcggtaaa gctcatcagc gtggtcgtga agcgattcac agatgtctgc
     3841 ctgttcatcc gcgtccagct cgttgagttt ctccagaagc gttaatgtct ggcttctgat
     3901 aaagcgggcc atgttaaggg cggttttttc ctgtttggtc actgatgcct ccgtgtaagg
     3961 gggatttctg ttcatggggg taatgatacc gatgaaacga gagaggatgc tcacgatacg
     4021 ggttactgat gatgaacatg cccggttact ggaacgttgt gagggtaaac aactggcggt
     4081 atggatgcgg cgggaccaga gaaaaatcac tcagggtcaa tgccagcgct tcgttaatac
     4141 agatgtaggt gttccacagg gtagccagca gcatcctgcg atgcagatcc ggaacataat
     4201 ggtgcagggc gctgacttcc gcgtttccag actttacgaa acacggaaac cgaagaccat
     4261 tcatgttgtt gctcaggtcg cagacgtttt gcagcagcag tcgcttcacg ttcgctcgcg
     4321 tatcggtgat tcattctgct aaccagtaag gcaaccccgc cagcctagcc gggtcctcaa
     4381 cgacaggagc acgatcatgc gcacccgtgg ggccgccatg ccggcgataa tggcctgctt
     4441 ctcgccgaaa cgtttggtgg cgggaccagt gacgaaggct tgagcgaggg cgtgcaagat
     4501 tccgaatacc gcaagcgaca ggccgatcat cgtcgcgctc cagcgaaagc ggtcctcgcc
     4561 gaaaatgacc cagagcgctg ccggcacctg tcctacgagt tgcatgataa agaagacagt
     4621 cataagtgcg gcgacgatag tcatgccccg cgcccaccgg aaggagctga ctgggttgaa
     4681 ggctctcaag ggcatcggtc gagatcccgg tgcctaatga gtgagctaac ttacattaat
     4741 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     4801 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgccagggtg gtttttcttt
     4861 tcaccagtga gacgggcaac agctgattgc ccttcaccgc ctggccctga gagagttgca
     4921 gcaagcggtc cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg
     4981 gcgggatata acatgagctg tcttcggtat cgtcgtatcc cactaccgag atatccgcac
     5041 caacgcgcag cccggactcg gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg
     5101 caaccagcat cgcagtggga acgatgccct cattcagcat ttgcatggtt tgttgaaaac
     5161 cggacatggc actccagtcg ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga
     5221 gatatttatg ccagccagcc agacgcagac gcgccgagac agaacttaat gggcccgcta
     5281 acagcgcgat ttgctggtga cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt
     5341 cttcatggga gaaaataata ctgttgatgg gtgtctggtc agagacatca agaaataacg
     5401 ccggaacatt agtgcaggca gcttccacag caatggcatc ctggtcatcc agcggatagt
     5461 taatgatcag cccactgacg cgttgcgcga gaagattgtg caccgccgct ttacaggctt
     5521 cgacgccgct tcgttctacc atcgacacca ccacgctggc acccagttga tcggcgcgag
     5581 atttaatcgc cgcgacaatt tgcgacggcg cgtgcagggc cagactggag gtggcaacgc
     5641 caatcagcaa cgactgtttg cccgccagtt gttgtgccac gcggttggga atgtaattca
     5701 gctccgccat cgccgcttcc actttttccc gcgttttcgc agaaacgtgg ctggcctggt
     5761 tcaccacgcg ggaaacggtc tgataagaga caccggcata ctctgcgaca tcgtataacg
     5821 ttactggttt cacattcacc accctgaatt gactctcttc cgggcgctat catgccatac
     5881 cgcgaaaggt tttgcgccat tcgatggtgt ccgggatctc gacgctctcc cttatgcgac
     5941 tcctgcatta ggaagcagcc cagtagtagg ttgaggccgt tgagcaccgc cgccgcaagg
     6001 aatggtgcat gcaaggagat ggcgcccaac agtcccccgg ccacggggcc tgccaccata
     6061 cccacgccga aacaagcgct catgagcccg aagtggcgag cccgatcttc cccatcggtg
     6121 atgtcggcga tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc ggccacgatg
     6181 cgtccggcgt agaggatcg
//
