LOCUS       sequence-257126-        5931 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expresses KSHV ORF18 (R158A_L159A) with Cterm 3xFLAG tag in 
            mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5931)
  AUTHORS   Castaneda AF, Glaunsinger BA
  TITLE     The interaction between ORF18 and ORF30 is required for late gene 
            expression in Kaposi's sarcoma-associated herpesvirus.
  JOURNAL   J Virol. 2018 Oct 10. pii: JVI.01488-18. doi: 10.1128/JVI.01488-18.
  PUBMED    30305361
REFERENCE   2  (bases 1 to 5931)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Virol. 
            2018 Oct 10. pii: JVI.01488-18. doi: 10.1128/JVI.01488-18."
COMMENT     Sequence Label: ORF18(R158A_L159A)-3xFLAG pCDNA4
FEATURES             Location/Qualifiers
     source          1..5931
                     /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"
     protein_bind    820..838
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    841..859
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     869..893
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             1848..1913
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(1939..1956)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1945..2169
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2215..2643
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2302..2321)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2512..2533
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2652..2672)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2657..2986
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2837..2972
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2899..2918
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3034..3081
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3100..3474
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISTVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    3604..3725
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3641..3660)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3695..3714
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3774..3790)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3774..3790)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3787..3809)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3798..3814
                     /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(3822..3828,3829..3846,3847..3852))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3867..3888
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4005..4022)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4176..4764)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4256..4275)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4935..5726,5727..5795))
                     /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     5558..5577
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5796..5900)
                     /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 ccctatcagt gatagagatc
      841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga
      901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga
      961 ctctagcgtt taaacttaag cttggtaccg agctcggatc cactagtcca gtgtggtgga
     1021 attctgcaga tatgctcgga aaatacgtgt gtgagaccga gccgctgagc ccggggctta
     1081 ggaggctcat gtggcgcttt ttgcaaaata agaatttaaa tacattccac gcccaagagc
     1141 tgcgttttat tcatttggtt ctctgcagga tgtacaattt cggtctaaat gtgtacctgt
     1201 taagggaggc tactgccaat gccgggacct acgacgaggt ggtcctggga cgcaaggttc
     1261 ctgcggaggt gtggaagctc gtgtacgatg ggctcgagga gatgggcgtg tcaagtgaga
     1321 tgctgctgtg tgaggcatac cgggacagcc tctggatgca cttgaacgat aaggtggggc
     1381 tcttgagggg cctggcgaat tatctgtttc accggctagg ggtcacccac gacgttcgca
     1441 tcgccccgga aaacctggtg gacggaaact ttttgtttaa tctgggaagt gtgctcccct
     1501 gcgcggcgct ccttgcggcg ggctactgcc tcgccttttg gggcagcgat gaacacgaac
     1561 gctgggtgcg cttcttcgcc cagaagcttt tcatttgcta cctgatagtc tccgggcgtc
     1621 ttatgacaca gaggtctctg ctagtttggg ccagcgaaac gggctatccc ggtccggtgg
     1681 aggcagtctg tcgcgacatc cgctccatgt acggcatacg aacgtatgcg gtctcgggtt
     1741 atcttccggc tccgtccgaa gcgcagctgg cctaccttgg tgcgtttaac aacaacgcgg
     1801 tttcagatcc agcacagtgg cggccgctcg agggaggcgg tggagccgac tacaaggacc
     1861 acgacggcga ctacaaggac cacgacatcg actacaagga cgacgacgac aaggggcccg
     1921 tttaaacccg ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc
     1981 cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa
     2041 atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg
     2101 ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg
     2161 gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat ccccacgcgc
     2221 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     2281 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     2341 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     2401 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc
     2461 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     2521 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     2581 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     2641 attaattctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg
     2701 cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg
     2761 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc
     2821 gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca
     2881 tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct ctgagctatt
     2941 ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tcccgggagc
     3001 ttgtatatcc attttcggat ctgatcagca cgtgttgaca attaatcatc ggcatagtat
     3061 atcggcatag tataatacga caaggtgagg aactaaacca tggccaagtt gaccagtgcc
     3121 gttccggtgc tcaccgcgcg cgacgtcgcc ggagcggtcg agttctggac cgaccggctc
     3181 gggttctccc gggacttcgt ggaggacgac ttcgccggtg tggtccggga cgacgtgacc
     3241 ctgttcatca gcacggtcca ggaccaggtg gtgccggaca acaccctggc ctgggtgtgg
     3301 gtgcgcggcc tggacgagct gtacgccgag tggtcggagg tcgtgtccac gaacttccgg
     3361 gacgcctccg ggccggccat gaccgagatc ggcgagcagc cgtgggggcg ggagttcgcc
     3421 ctgcgcgacc cggccggcaa ctgcgtgcac ttcgtggccg aggagcagga ctgacacgtg
     3481 ctacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc
     3541 cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac
     3601 cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     3661 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     3721 tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag
     3781 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     3841 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     3901 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     3961 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     4021 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     4081 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     4141 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     4201 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     4261 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     4321 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     4381 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     4441 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     4501 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     4561 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     4621 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     4681 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     4741 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     4801 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     4861 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     4921 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     4981 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     5041 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     5101 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     5161 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     5221 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     5281 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     5341 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     5401 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     5461 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     5521 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     5581 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     5641 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     5701 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     5761 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     5821 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     5881 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt c
//
