LOCUS       sequence-243726-        6667 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Nuclear localised mRFP reporter construct with one half of split 
            Venus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6667)
  TITLE     Schirmer lab plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6667)
  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: 
            "Unpublished"
COMMENT     Sequence Label: NLS-mRFP-V1
FEATURES             Location/Qualifiers
     source          1..6667
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        116..495
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        496..699
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     650..670
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     744..763
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        744..762
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             842..862
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             869..1543
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /note="mammalian codon-optimized"
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVIQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     primer_bind     complement(999..1017)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1461..1484
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     CDS             join(1592..1594,1595..2053)
                     /codon_start=1
                     /product="improved N-terminal fragment of mVenus for use in
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN155(I152L)
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     LICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITA"
     primer_bind     complement(1634..1655)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1895..1914)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     promoter        complement(2101..2119)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2102..2119)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2139..2156)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2145..2369
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2415..2843
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2502..2521)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2712..2733
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2852..2872)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2857..3186
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3037..3172
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3099..3118
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3253..4047
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3307..3326)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3917..3936
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4221..4342
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4258..4277)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4312..4331
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4391..4407)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4391..4407)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4404..4426)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4415..4431
                     /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(4439..4445,4446..4463,4464..4469))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4484..4505
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4622..4639)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4793..5381)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4873..4892)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5552..6343,6344..6412))
                     /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     6175..6194
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6413..6517)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6592..6611)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat gaagaatctg
       61 cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac gcgttgacat
      121 tgattattga ctagttatta atagtaatca attatggggt cattagttca tagcccatat
      181 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
      241 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
      301 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
      361 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
      421 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
      481 atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt
      541 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
      601 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc
      661 ggtaggcgtg tacggtggga ggtctatata agcagagctc tctggctaac tagagaaccc
      721 actgcttact ggcttatcga aattaatacg actcactata gggagaccca agcttggtac
      781 cgagctcgga tccactagta acggccgcca gtgtgctgga attcctgcag gcctcgagat
      841 gcccaagaag aagaggaagg tgatggccat ggcctcctcc gaggacgtca tcaaggagtt
      901 catgcgcttc aaggtgcgca tggagggctc cgtgaacggc cacgagttcg agatcgaggg
      961 cgagggcgag ggccgcccct acgagggcac ccagaccgcc aagctgaagg tgaccaaggg
     1021 cggccccctg cccttcgcct gggacatcct gtcccctcag ttccagtacg gctccaaggc
     1081 ctacgtgaag caccccgccg acatccccga ctacttgaag ctgtccttcc ccgagggctt
     1141 caagtgggag cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga cccaggactc
     1201 ctccctgcag gacggcgagt tcatctacaa ggtgaagctg cgcggcacca acttcccctc
     1261 cgacggcccc gtaattcaga agaagacgat gggctgggag gcctccaccg agcggatgta
     1321 ccccgaggac ggcgccctga agggcgagat caagatgagg ctgaagctga aggacggcgg
     1381 ccactacgac gccgaggtca agaccaccta catggccaag aagcccgtgc agctgcccgg
     1441 cgcctacaag accgacatca agctggacat cacctcccac aacgaggact acaccatcgt
     1501 ggaacagtac gagcgcgccg agggccgcca ctccaccggc gcctctagag gatccggtgg
     1561 cggtggctct ggaggtggtg ggtcctccgg agtgagcaag ggcgaggagc tgttcaccgg
     1621 ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc
     1681 cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagctga tctgcaccac
     1741 cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgggctacg gcctgcagtg
     1801 cttcgcccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga
     1861 aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc
     1921 cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt
     1981 caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca gccacaacgt
     2041 ctatatcacc gccgacaagc agtaagcggc cgcgtcgagc atgcatctag agggccctat
     2101 tctatagtgt cacctaaatg ctagagctcg ctgatcagcc tcgactgtgc cttctagttg
     2161 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     2221 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
     2281 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag
     2341 gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc
     2401 tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     2461 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     2521 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     2581 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     2641 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     2701 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     2761 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
     2821 ttaacaaaaa tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag
     2881 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     2941 aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     3001 tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt
     3061 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc
     3121 gcctctgcct ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt
     3181 tgcaaaaagc tcccgggagc ttgtatatcc attttcggat ctgatcaaga gacaggatga
     3241 ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg
     3301 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg
     3361 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc
     3421 ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct
     3481 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa
     3541 gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg
     3601 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa
     3661 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat
     3721 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg
     3781 cgcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc
     3841 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac
     3901 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg
     3961 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc
     4021 tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag
     4081 cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg
     4141 gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc
     4201 tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca
     4261 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     4321 ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg
     4381 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     4441 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     4501 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     4561 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     4621 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     4681 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     4741 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     4801 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     4861 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     4921 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     4981 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     5041 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     5101 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     5161 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     5221 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     5281 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     5341 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     5401 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     5461 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     5521 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     5581 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     5641 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     5701 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     5761 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     5821 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     5881 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     5941 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     6001 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     6061 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     6121 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     6181 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     6241 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     6301 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     6361 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     6421 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     6481 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     6541 ctgacgtcga cggatcggga gatctcccga tcccctatgg tgcactctca gtacaatctg
     6601 ctctgatgcc gcatagttaa gccagtatct gctccctgct tgtgtgttgg aggtcgctga
     6661 gtagtgc
//
