LOCUS       sequence-345721-        6719 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of Flag-tagged KSHV ORF45 RC mutant.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6719)
  AUTHORS   Alzhanova D, Meyo JO, Juarez A, Dittmer DP
  TITLE     The ORF45 Protein of Kaposi Sarcoma-Associated Herpesvirus Is an 
            Inhibitor of p53 Signaling during Viral Reactivation.
  JOURNAL   J Virol. 2021 Nov 9;95(23):e0145921. doi: 10.1128/JVI.01459-21. Epub
            2021 Sep 15.
  PUBMED    34523970
REFERENCE   2  (bases 1 to 6719)
  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: "J Virol."; 
            date: "2021-11-9"; volume: "95(23):e0145921. doi"; pages: " 
            10.1128/JVI.01459-21. Epub 2021 Sep 15"
COMMENT     Sequence Label: pDD2706
FEATURES             Location/Qualifiers
     source          1..6719
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      944..953
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2171..2236
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     promoter        complement(2274..2292)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2275..2292)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2312..2329)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2318..2542
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2588..3016
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2675..2694)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2885..2906
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3025..3045)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3030..3359
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3210..3345
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3272..3291
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     complement(3480..3499)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4088..4107
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4392..4513
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4429..4448)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4483..4502
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4562..4578)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4562..4578)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4575..4597)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4586..4602
                     /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)."
     promoter        complement(join(4610..4616,4617..4634,4635..4640))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4655..4676
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4793..4810)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4964..5552)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5044..5063)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5723..6514,6515..6583))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6346..6365
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6584..6688)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttggtacc
      901 gagctcggat ccactagtaa cggccgccag tgtgctggaa ttcgccacca tggcgatgtt
      961 tgtgaggacc tcgtctagca cacacgatga agagagaatg cttccaattg aaggagcgcc
     1021 tcgcagacga ccccccgtga agttcatatt cccacctcca cctctttcat cacttccagg
     1081 atttggcagg ccgcgcggct atgctggacc cacggtgata gatatgtctg ccccagacga
     1141 cgtcttcgcc gaggacacgc catcgccgcc agcaacccct ctggatctac agatatcccc
     1201 ggatcagtcg agcggcgaat ctgaatatga cgaggatgag gaagatgaag atgaagaaga
     1261 aaatgacgat gttcaggagg aagacgagcc agaggggtac cctgcagact tttttcaacc
     1321 tttatctcac ttgcgcccga ggcctctggc cagacgggcc catacgccca aaccggtagc
     1381 agtggtagcg ggccgcgtgc gcagttcaac ggacacggcg gagtccgagg cgtccatggg
     1441 atgggttagt caggatgacg gattttcccc tgctgggctc tcaccttcag acgacgaggg
     1501 ggttgctatc ctggaaccga tggcggcata cactgggacc ggggcatacg gactttcacc
     1561 tgcttccaga aatagtgtac ctggaacaca aagttcacca tacagcgacc ctgatgaagg
     1621 gccctcgtgg cgccccctgc gcgccgcacc caccgcgatc gtcgacctga catcggactc
     1681 tgatagcgat gacagttcca actctccgga cgtgaacaat gaggccgcgt ttaccgacgc
     1741 gcgccatttt tcccaccagc caccctcgtc cgaggaggac ggagaagacc aaggggaagt
     1801 attgagtcag agaatcgggc tcatggacgt gggccaggcg gccgcagcgc agtctaccgc
     1861 ctcctctggt agcgaggatg tggtgcgctg ccagagacaa ccaaacttaa gccgcaaagc
     1921 agtggcgtcc gtgataatta tatcctcggg gagtgacaca gacgaggagc cctcgtccgc
     1981 cgtgagcgtg atcgtgtctc cgtcgagcac aaagggtcac ctcccaaccc aatctcccag
     2041 tacttccgcc cactcgattt catcaggaag cacaactacc gcggggtcca ggtgcagcga
     2101 cccaacccgc atcctggcct ccacgccacc cctgtgtgga aacggtgcat ataactggcc
     2161 gtggctggac gactacaagg accacgacgg tgactacaag gaccacgaca tcgactacaa
     2221 ggacgacgac gacaagtgag cggccgctcg agcatgcatc tagagggccc tattctatag
     2281 tgtcacctaa atgctagagc tcgctgatca gcctcgactg tgccttctag ttgccagcca
     2341 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
     2401 ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg
     2461 gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct
     2521 ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg
     2581 tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     2641 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     2701 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     2761 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     2821 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     2881 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     2941 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     3001 aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
     3061 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
     3121 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     3181 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     3241 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg
     3301 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
     3361 agctcccggg agcttgtata tccattttcg gatctgatca agagacagga tgaggatcgt
     3421 ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc
     3481 tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc
     3541 tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg
     3601 aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag
     3661 ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg
     3721 ggcaggatct cctcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca
     3781 atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat
     3841 cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac
     3901 gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc
     3961 gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa
     4021 aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag
     4081 gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc
     4141 ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt
     4201 cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca
     4261 acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa
     4321 tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct
     4381 tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     4441 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     4501 tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat
     4561 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     4621 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     4681 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     4741 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     4801 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     4861 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     4921 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     4981 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     5041 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     5101 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     5161 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     5221 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     5281 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     5341 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     5401 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     5461 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     5521 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     5581 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     5641 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     5701 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     5761 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     5821 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     5881 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     5941 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     6001 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     6061 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     6121 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     6181 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     6241 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     6301 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     6361 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     6421 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     6481 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     6541 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     6601 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     6661 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtc
//
