LOCUS       sequence-261677-        7100 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expresses Flag-tagged HROB in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7100)
  AUTHORS   Hustedt N, Saito Y, Zimmermann M, Alvarez-Quilon A, Setiaputra D, 
            Adam S, McEwan A, Yuan JY, Olivieri M, Zhao Y, Kanemaki MT, 
            Jurisicova A, Durocher D
  TITLE     Control of homologous recombination by the HROB-MCM8-MCM9 pathway.
  JOURNAL   Genes Dev. 2019 Oct 1;33(19-20):1397-1415. doi: 
            10.1101/gad.329508.119. Epub 2019 Aug 29.
  PUBMED    31467087
REFERENCE   2  (bases 1 to 7100)
  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: "Genes 
            Dev."; date: "2019-10-1"; volume: "33(19-20):1397-1415. doi"; pages:
            " 10.1101/gad.329508.119. Epub 2019 Aug 29"
COMMENT     Sequence Label: pcDNA5-FRT-T0-Flag-HROB
FEATURES             Location/Qualifiers
     source          1..7100
                     /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             993..1016
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(3052..3069)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3058..3282
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3415..3434)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3566..3613
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             3621..4643
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGDFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    4773..4894
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4810..4829)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4864..4883
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4943..4959)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4943..4959)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4956..4978)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4967..4983
                     /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(4991..4997,4998..5015,5016..5021))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5036..5057
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5174..5191)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5345..5933)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5425..5444)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6104..6895,6896..6964))
                     /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     6727..6746
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6965..7069)
                     /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 cttggtacca tggactacaa agacgatgac gataaagccg
     1021 ccagtttaaa cggcgcgcca gcgtgcagtt tgcagaagct gtttgctgtg gaagaggagt
     1081 ttgaagatga ggatttcttg tctgctgtgg aggatgcaga gaaccggttt actggctcac
     1141 tgcctgtgaa tgctgggcgc ctgagacctg tctcttctag gccacaggag actgtgcagg
     1201 cacagtcctc caggctgctg ctgttacacc ccactgctcc ctcagaggct ttgggcctgc
     1261 cagacttgga cctctgcctc cctgcctcca gcacgcccag tgctgacagc cgtccatcat
     1321 gcataggagc agctccccta aggcctgtct ctacttccag cagctggatt ggcaatcaga
     1381 gaagagtgac agtgacagaa gtgctcagag agacagcaag acctcagtcc tcagccttac
     1441 accccctact cacctttgag agccaacagc agcaagttgg tggctttgag gggcctgaac
     1501 aagacgaatt tgataaagtc ctggcaagca tggagttgga ggagcctggc atggagctgg
     1561 aatgtggagt cagcagtgag gccataccaa tcctgcctgc ccagcagcgg gagggttcag
     1621 tattggctaa aaaagcccgg gtagttgatc tgagtggatc ttgccagaag gggcctgtgc
     1681 ctgccatcca caaagcgggt atcatgtccg cccaggatga gtctctagat cctgtcatcc
     1741 aatgtaggac tccacgaccc cccttgagac ctggtgctgt gggtcacctt cctgttccaa
     1801 ctgccttaac agttcccact cagcaactcc actgggaagt ctgtccgcaa cgctcccctg
     1861 ttcaagcact tcagcctctc caagctgcta gagggaccat tcagagcagc cctcaaaatc
     1921 gtttcccttg tcagccattc cagtctccaa gttcctggtt aagtggcaaa gctcatttac
     1981 ccagacctcg aactcccaac tcaagctgtt ctactccctc aaggactagc tctggattat
     2041 ttcctcggat acccttacaa ccgcaagctc cagtgtcttc cattgggtct cctgttggta
     2101 ccccaaaagg tccccaggga gctctgcaga cacccatagt caccaaccac ctggtgcagc
     2161 tagtcactgc tgccagccgg acaccccagc agcccaccca tccctccacc cgagccaaaa
     2221 ctcgccgttt ccctggccca gctgggatcc tgcctcacca gcagagtggg agaagtctgg
     2281 aggacatcat ggtttccgcg ccccaaactc caacccatgg tgctctggct aaattccaga
     2341 cagagattgt tgctagttcc caggcatctg tggaggagga ttttgggcga gggccctggc
     2401 tgaccatgaa atccacgcta ggcctggatg agagagaccc tagctgcttc ctctgtacct
     2461 acagcattgt catggtgctg cgcaagcagg cagccctgaa gcagcttcct aggaacaagg
     2521 tccccaacat ggcggtgatg atcaagtccc tgactcggag cacaatggac gccagtgtgg
     2581 ttttcaagga ccccacggga gagatgcagg ggacggtgca caggttgctg ctggagacgt
     2641 gccagaatga gctgaagcct ggctcagtgc tgctgctgaa gcagattgga gtgttttctc
     2701 cttcacttcg aaatcactac ctcaacgtga cacccaacaa cctggtccat atttacagcc
     2761 cggattctgg ggatgggagc ttcctcaagc catctcagcc cttccccaag gattcaggga
     2821 gcttccagca tgatgtggct gcaaagcccg aggaaggctt cagaacagca cagaacctag
     2881 aggcagaggc gtcccctgag gaagaactcc cagaagcaga tgacctggat ggactcctga
     2941 gtgagcttcc tgaagacttc ttctgtggga ccagtagttg agaattctgc agatatccat
     3001 cacactggcg gccgctcgag tctagagggc ccgtttaaac ccgctgatca gcctcgactg
     3061 tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg
     3121 aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga
     3181 gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg
     3241 aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa
     3301 ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg
     3361 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     3421 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     3481 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     3541 attagggtga tggttcacgt acctagaagt tcctattccg aagttcctat tctctagaaa
     3601 gtataggaac ttccttggcc aaaaagcctg aactcaccgc gacgtctgtc gagaagtttc
     3661 tgatcgaaaa gttcgacagc gtctccgacc tgatgcagct ctcggagggc gaagaatctc
     3721 gtgctttcag cttcgatgta ggagggcgtg gatatgtcct gcgggtaaat agctgcgccg
     3781 atggtttcta caaagatcgt tatgtttatc ggcactttgc atcggccgcg ctcccgattc
     3841 cggaagtgct tgacattggg gatttcagcg agagcctgac ctattgcatc tcccgccgtg
     3901 cacagggtgt cacgttgcaa gacctgcctg aaaccgaact gcccgctgtt ctgcagccgg
     3961 tcgcggaggc catggatgcg atcgctgcgg ccgatcttag ccagacgagc gggttcggcc
     4021 cattcggacc gcaaggaatc ggtcaataca ctacatggcg tgatttcata tgcgcgattg
     4081 ctgatcccca tgtgtatcac tggcaaactg tgatggacga caccgtcagt gcgtccgtcg
     4141 cgcaggctct cgatgagctg atgctttggg ccgaggactg ccccgaagtc cggcacctcg
     4201 tgcacgcgga tttcggctcc aacaatgtcc tgacggacaa tggccgcata acagcggtca
     4261 ttgactggag cgaggcgatg ttcggggatt cccaatacga ggtcgccaac atcttcttct
     4321 ggaggccgtg gttggcttgt atggagcagc agacgcgcta cttcgagcgg aggcatccgg
     4381 agcttgcagg atcgccgcgg ctccgggcgt atatgctccg cattggtctt gaccaactct
     4441 atcagagctt ggttgacggc aatttcgatg atgcagcttg ggcgcagggt cgatgcgacg
     4501 caatcgtccg atccggagcc gggactgtcg ggcgtacaca aatcgcccgc agaagcgcgg
     4561 ccgtctggac cgatggctgt gtagaagtac tcgccgatag tggaaaccga cgccccagca
     4621 ctcgtccgag ggcaaaggaa tagcacgtac tacgagattt cgattccacc gccgccttct
     4681 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
     4741 gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt
     4801 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     4861 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta
     4921 gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     4981 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     5041 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     5101 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     5161 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     5221 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     5281 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     5341 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     5401 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     5461 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     5521 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     5581 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     5641 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     5701 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     5761 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     5821 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     5881 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     5941 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     6001 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     6061 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     6121 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     6181 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     6241 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     6301 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     6361 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     6421 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     6481 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     6541 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     6601 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     6661 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     6721 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     6781 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     6841 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     6901 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     6961 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     7021 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     7081 gaaaagtgcc acctgacgtc
//
