LOCUS       sequence-304979-        6743 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Natively produces mNeonGreen, in the presence of Flp or Flpe will be
            recombined to produce mScarlet..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6743)
  AUTHORS   Taylor RJ, Carrington J, Gerlach LR, Taylor KL, Richters KE, Dent EW
  TITLE     Double UP: A Dual Color, Internally Controlled Platform for in utero
            Knockdown or Overexpression.
  JOURNAL   Front Mol Neurosci. 2020 May 20;13:82. doi: 
            10.3389/fnmol.2020.00082. eCollection 2020.
  PUBMED    32508591
REFERENCE   2  (bases 1 to 6743)
  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: "Front Mol 
            Neurosci."; date: "2020-05-20"; volume: "13:82. doi"; pages: " 
            10.3389/fnmol.2020.00082. eCollection 2020"
COMMENT     Sequence Label: Double UP mNeon to mScarlet FRT
FEATURES             Location/Qualifiers
     source          1..6743
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        5..384
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          662..1678
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1602..1624
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1686..1705
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    1739..1772
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     CDS             1779..2489
                     /codon_start=1
                     /product="bright monomeric yellow-green fluorescent protein
                     derived from LanYFP (Shaner et al., 2013)"
                     /label=mNeonGreen
                     /translation="MVSKGEEDNMASLPATHELHIFGSINGVDFDMVGQGTGNPNDGYE
                     ELNLKSTKGDLQFSPWILVPHIGYGFHQYLPYPDGMSPFQAAMVDGSGYQVHRTMQFED
                     GASLTVNYRYTYEGSHIKGEAQVKGTGFPADGPVMTNSLTAADWCRSKKTYPNDKTIIS
                     TFKWSYTTGNGKRYRSTARTTYTFAKPMAANYLKNQPMYVFRKTELKHSKTELNFKEWQ
                     KAFTDVMGMDELYK"
     regulatory      2034..2043
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2558..2613
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     protein_bind    2929..2962
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     CDS             2969..3664
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet-I
                     /note="matures faster than mScarlet"
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFIKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(3102..3120)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3370..3389
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             3680..3736
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     polyA_signal    3839..3894
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4246..4262)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4246..4262)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4259..4281)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4270..4286
                     /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(4294..4300,4301..4318,4319..4324))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4339..4360
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4419..4614
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4465..4600
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4620..4754
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4657..4676)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4711..4730
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4822..4839)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4993..5581)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5073..5092)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5752..6543,6544..6612))
                     /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     6375..6394
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6613..6717)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
       61 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      121 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      181 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      241 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      301 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      361 gtattagtca tcgctattac catggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggggcgc gccrgggggg gggggggggg gggggggggg
      541 gggsggggsg rggsggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgcgctgcc ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc
      721 gccccggctc tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc
      781 tccgggctgt aattagcgct tggtttaatg acggcttgtt tcttttctgt ggctgcgtga
      841 aagccttgag gggctccggg agggcccttt gtgcgggggg agcggctcgg ggggtgcgtg
      901 cgtgtgtgtg tgcgtgggga gcgccgcgtg cggctccgcg ctgcccggcg gctgtgagcg
      961 ctgcgggcgc ggcgcggggc tttgtgcgct ccgcagtgtg cgcgagggga gcgcggccgg
     1021 gggcggtgcc ccgcggtgcg gggggggctg cgaggggaac aaaggctgcg tgcggggtgt
     1081 gtgcgtgggg gggtgagcag ggggtgtggg cgcgtcggtc gggctgcaac cccccctgca
     1141 cccccctccc cgagttgctg agcacggccc ggcttcgggt gcggggctcc gtacggggcg
     1201 tggcgcgggg ctcgccgtgc cgggcggggg gtggcggcag gtgggggtgc cgggcggggc
     1261 ggggccgcct cgggccgggg rgggctcggg ggaggggcgc ggcggccccc ggagcgccgg
     1321 cggctgtcga ggcgcggcga gccgcagcca ttgcctttta tggtaatcgt gcgagagggc
     1381 gcagggactt cctttgtccc aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac
     1441 cccctctagc gggcgcgggg cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga
     1501 gggccttcgt gcgtcgccgc gccgccgtcc ccttctccct ctccagcctc ggggctgtcc
     1561 gcggggggac ggctgccttc gggggggacg gggcagggcg gggttcggct tctggcgtgt
     1621 gaccggcggc tctagagcct ctgctaacca tgttcatgcc ttcttctttt tcctacagct
     1681 cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt ggcaaagaat tgctgcagga
     1741 agttcctatt ctctagaaag tataggaact tcacgcgtat ggtgagcaag ggcgaggagg
     1801 ataacatggc ctctctccca gcgacacatg agttacacat ctttggctcc atcaacggtg
     1861 tggactttga catggtgggt cagggcaccg gcaatccaaa tgatggttat gaggagttaa
     1921 acctgaagtc caccaagggt gacctccagt tctccccctg gattctggtc cctcatatcg
     1981 ggtatggctt ccatcagtac ctgccctacc ctgacgggat gtcgcctttc caggccgcca
     2041 tggtagatgg ctccggctac caagtccatc gcacaatgca gtttgaagat ggtgcctccc
     2101 ttactgttaa ctaccgctac acctacgagg gaagccacat caaaggagag gcccaggtga
     2161 aggggactgg tttccctgct gacggtcctg tgatgaccaa ctcgctgacc gctgcggact
     2221 ggtgcaggtc gaagaagact taccccaacg acaaaaccat catcagtacc tttaagtgga
     2281 gttacaccac tggaaatggc aagcgctacc ggagcactgc gcggaccacc tacacctttg
     2341 ccaagccaat ggcggctaac tatctgaaga accagccgat gtacgtgttc cgtaagacgg
     2401 agctcaagca ctccaagacc gagctcaact tcaaggagtg gcaaaaggcc tttaccgatg
     2461 tgatgggcat ggacgagctg tacaagtaaa gatctttttc cctctgccaa aaattatggg
     2521 gacatcatga agccccttga gcatctgact tctggctaat aaaggaaatt tattttcatt
     2581 gcaatagtgt gttggaattt tttgtgtctc tcactcggaa ggacatatgg gagggcaaat
     2641 catttaaaac atcagaatga gtatttggtt tagagtttgg caacatatgc ccatatgctg
     2701 gctgccatga acaaaggttg gctataaaga ggtcatcagt atatgaaaca gccccctgct
     2761 gtccattcct tattccatag aaaagccttg acttgaggtt agattttttt tatattttgt
     2821 tttgtgttat ttttttcttt aacatcccta aaattttcct tacatgtttt actagccaga
     2881 tttttcctcc tctcctgact actcccagtc atagctgtcc ctcttctcga agttcctatt
     2941 ctctagaaag tataggaact tcggtaccat ggtgagcaag ggcgaggcag tgatcaagga
     3001 gttcatgcgg ttcaaggtgc acatggaggg ctccatgaac ggccacgagt tcgagatcga
     3061 gggcgagggc gagggccgcc cctacgaggg cacccagacc gccaagctga aggtgaccaa
     3121 gggtggcccc ctgcccttct cctgggacat cctgtcccct cagttcatgt acggctccag
     3181 ggccttcatc aagcaccccg ccgacatccc cgactactat aagcagtcct tccccgaggg
     3241 cttcaagtgg gagcgcgtga tgaacttcga ggacggcggc gccgtgaccg tgacccagga
     3301 cacctccctg gaggacggca ccctgatcta caaggtgaag ctccgcggca ccaacttccc
     3361 tcctgacggc cccgtaatgc agaagaagac aatgggctgg gaagcgtcca ccgagcggtt
     3421 gtaccccgag gacggcgtgc tgaagggcga cattaagatg gccctgcgcc tgaaggacgg
     3481 cggacgctac ctggcggact tcaagaccac ctacaaggcc aagaagcccg tgcagatgcc
     3541 cggcgcctac aacgtcgacc gcaagttgga catcacctcc cacaacgagg actacaccgt
     3601 ggtggaacag tacgaacgct ccgagggccg ccactccacc ggcggcatgg acgagctgta
     3661 caagttcgaa ggcagcggcg ctactaactt cagcctgctg aagcaggctg gagacgtgga
     3721 agagaatcct ggacctatcg ataccggtca attggcggcc gcgcatgcgc tagctaattt
     3781 ccctctgcca aaaattatgg ggacatcatg aagccccttg agcatctgac ttctggctaa
     3841 taaaggaaat ttattttcat tgcaatagtg tgttggaatt ttttgtgtct ctcactcgga
     3901 aggacatatg ggagggcaaa tcatttaaaa catcagaatg agtatttggt ttagagtttg
     3961 gcaacatatg cccatatgct ggctgccatg aacaaaggtt ggctataaag aggtcatcag
     4021 tatatgaaac agccccctgc tgtccattcc ttattccata gaaaagcctt gacttgaggt
     4081 tagatttttt ttatattttg ttttgtgtta tttttttctt taacatccct aaaattttcc
     4141 ttacatgttt tactagccag atttttcctc ctctcctgac tactcccagt catagctgtc
     4201 cctcttctct tatggagatc cctcgacaag cttggcgtaa tcatggtcat agctgtttcc
     4261 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     4321 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     4381 cgctttccag tcgggaaacc tgtcgtgcca gcggatccgc atctcaatta gtcagcaacc
     4441 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     4501 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct
     4561 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagcta
     4621 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     4681 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     4741 atcatgtctg gatccgctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     4801 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     4861 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     4921 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     4981 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     5041 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     5101 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     5161 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     5221 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     5281 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     5341 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     5401 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     5461 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     5521 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     5581 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     5641 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     5701 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     5761 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     5821 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     5881 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     5941 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     6001 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     6061 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     6121 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     6181 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     6241 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     6301 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     6361 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     6421 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     6481 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     6541 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     6601 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     6661 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     6721 atttccccga aaagtgccac ctg
//
