LOCUS       sequence-416611-        5847 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  N-terminal mCherry-tagging of inserts for inducible expression. Use 
            with Flp-In T-Rex system..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5847)
  AUTHORS   Hertz EPT, Vega IA, Kruse T, Wang Y, Hendriks IA, Bizard AH, 
            Eugui-Anta A, Hay RT, Nielsen ML, Nilsson J, Hickson ID, Mailand N
  TITLE     The SUMO-NIP45 pathway processes toxic DNA catenanes to prevent 
            mitotic failure.
  JOURNAL   Nat Struct Mol Biol. 2023 Jul 20. doi: 10.1038/s41594-023-01045-0.
  PUBMED    37474739
REFERENCE   2  (bases 1 to 5847)
  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: "Nat Struct 
            Mol Biol. 2023 Jul 20. doi: 10.1038/s41594-023-01045-0."
COMMENT     Sequence Label: pcDNA5-FRT-TO-mCherry-N
FEATURES             Location/Qualifiers
     source          1..5847
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     23..47
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      134..143
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             140..847
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(285..303)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     553..572
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(953..970)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    959..1183
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(1316..1335)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    1467..1514
                     /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             1522..2544
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             1522..2544
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    2674..2795
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2711..2730)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2765..2784
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2844..2860)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2844..2860)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2857..2879)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2868..2884
                     /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(2892..2898,2899..2916,2917..2922))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2937..2958
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3075..3092)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3246..3834)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3326..3345)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4005..4796,4797..4865))
                     /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     4628..4647
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4866..4970)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5045..5064)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        5236..5615
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5616..5819
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5770..5790
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    5821..5839
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    join(5842..5847,1..13)
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
ORIGIN
        1 tcagtgatag agatcgtcga cgagctcgtt tagtgaaccg tcagatcgcc tggagacgcc
       61 atccacgctg ttttgacctc catagaagac accgggaccg atccagcctc cggactctag
      121 cgtttaaact taagccacca tggtgagcaa gggcgaggag gataacatgg ccatcatcaa
      181 ggagttcatg cgcttcaagg tgcacatgga gggctccgtg aacggccacg agttcgagat
      241 cgagggcgag ggcgagggcc gcccctacga gggcacccag accgccaagc tgaaggtgac
      301 caagggtggc cccctgccct tcgcctggga catcctgtcc cctcagttca tgtacggctc
      361 caaggcctac gtgaagcacc ccgccgacat ccccgactac ttgaagctgt ccttccccga
      421 gggcttcaag tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca
      481 ggactcctcc ctgcaggacg gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt
      541 cccctccgac ggccccgtaa tgcagaagaa gaccatgggc tgggaggcct cctccgagcg
      601 gatgtacccc gaggacggcg ccctgaaggg cgagatcaag cagaggctga agctgaagga
      661 cggcggccac tacgacgctg aggtcaagac cacctacaag gccaagaagc ccgtgcagct
      721 gcccggcgcc tacaacgtca acatcaagtt ggacatcacc tcccacaacg aggactacac
      781 catcgtggaa cagtacgaac gcgccgaggg ccgccactcc accggcggca tggacgagct
      841 gtacaagggt accgagctcg gatccactag tccagtgtgg tggaattctg cagatatcca
      901 gcacagtggc ggccgctcga gtctagaggg cccgtttaaa cccgctgatc agcctcgact
      961 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg
     1021 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg
     1081 agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg
     1141 gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga
     1201 accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg
     1261 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct
     1321 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
     1381 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
     1441 gattagggtg atggttcacg tacctagaag ttcctattcc gaagttccta ttctctagaa
     1501 agtataggaa cttccttggc caaaaagcct gaactcaccg cgacgtctgt cgagaagttt
     1561 ctgatcgaaa agttcgacag cgtctccgac ctgatgcagc tctcggaggg cgaagaatct
     1621 cgtgctttca gcttcgatgt aggagggcgt ggatatgtcc tgcgggtaaa tagctgcgcc
     1681 gatggtttct acaaagatcg ttatgtttat cggcactttg catcggccgc gctcccgatt
     1741 ccggaagtgc ttgacattgg ggaattcagc gagagcctga cctattgcat ctcccgccgt
     1801 gcacagggtg tcacgttgca agacctgcct gaaaccgaac tgcccgctgt tctgcagccg
     1861 gtcgcggagg ccatggatgc gatcgctgcg gccgatctta gccagacgag cgggttcggc
     1921 ccattcggac cgcaaggaat cggtcaatac actacatggc gtgatttcat atgcgcgatt
     1981 gctgatcccc atgtgtatca ctggcaaact gtgatggacg acaccgtcag tgcgtccgtc
     2041 gcgcaggctc tcgatgagct gatgctttgg gccgaggact gccccgaagt ccggcacctc
     2101 gtgcacgcgg atttcggctc caacaatgtc ctgacggaca atggccgcat aacagcggtc
     2161 attgactgga gcgaggcgat gttcggggat tcccaatacg aggtcgccaa catcttcttc
     2221 tggaggccgt ggttggcttg tatggagcag cagacgcgct acttcgagcg gaggcatccg
     2281 gagcttgcag gatcgccgcg gctccgggcg tatatgctcc gcattggtct tgaccaactc
     2341 tatcagagct tggttgacgg caatttcgat gatgcagctt gggcgcaggg tcgatgcgac
     2401 gcaatcgtcc gatccggagc cgggactgtc gggcgtacac aaatcgcccg cagaagcgcg
     2461 gccgtctgga ccgatggctg tgtagaagta ctcgccgata gtggaaaccg acgccccagc
     2521 actcgtccga gggcaaagga atagcacgta ctacgagatt tcgattccac cgccgccttc
     2581 tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc
     2641 ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt
     2701 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     2761 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct
     2821 agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     2881 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     2941 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     3001 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     3061 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     3121 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     3181 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     3241 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     3301 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     3361 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     3421 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     3481 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     3541 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     3601 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     3661 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     3721 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     3781 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     3841 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     3901 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     3961 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     4021 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     4081 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     4141 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     4201 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     4261 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     4321 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     4381 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     4441 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     4501 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     4561 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     4621 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     4681 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     4741 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     4801 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     4861 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     4921 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     4981 cgaaaagtgc cacctgacgt cgacggatcg ggagatctcc cgatccccta tggtgcactc
     5041 tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctccct gcttgtgtgt
     5101 tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg
     5161 acaattgcat gaagaatctg cttagggtta ggcgttttgc gctgcttcgc gatgtacggg
     5221 ccagatatac gcgttgacat tgattattga ctagttatta atagtaatca attacggggt
     5281 cattagttca tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc
     5341 ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag
     5401 taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc
     5461 acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg
     5521 gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc
     5581 agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca
     5641 atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca
     5701 atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg
     5761 ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctc
     5821 tccctatcag tgatagagat ctcccta
//
