LOCUS       sequence-244757-        6737 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Natively produces mNeon, in the presence of Cre will be recombined 
            to produce mScarlet..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6737)
  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 6737)
  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
FEATURES             Location/Qualifiers
     source          1..6737
                     /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=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             1773..2483
                     /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      2028..2037
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2552..2607
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     protein_bind    2923..2956
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             2963..3658
                     /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(3096..3114)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3364..3383
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             3674..3730
                     /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    3833..3888
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4240..4256)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4240..4256)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4253..4275)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4264..4280
                     /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(4288..4294,4295..4312,4313..4318))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4333..4354
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4413..4608
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4459..4594
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4614..4748
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4651..4670)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4705..4724
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4816..4833)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4987..5575)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5067..5086)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5746..6537,6538..6606))
                     /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     6369..6388
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6607..6711)
                     /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 gggggsgcgc gccrggsggg gsggggsggg gsgrggggsg
      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 ggggccgccy cgggccgggg rgggctcggg ggrggggcgc 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 tgctgcagat
     1741 aacttcgtat aatgtatgct atacgaagtt atatggtgag caagggcgag gaggataaca
     1801 tggcctctct cccagcgaca catgagttac acatctttgg ctccatcaac ggtgtggact
     1861 ttgacatggt gggtcagggc accggcaatc caaatgatgg ttatgaggag ttaaacctga
     1921 agtccaccaa gggtgacctc cagttctccc cctggattct ggtccctcat atcgggtatg
     1981 gcttccatca gtacctgccc taccctgacg ggatgtcgcc tttccaggcc gccatggtag
     2041 atggctccgg ctaccaagtc catcgcacaa tgcagtttga agatggtgcc tcccttactg
     2101 ttaactaccg ctacacctac gagggaagcc acatcaaagg agaggcccag gtgaagggga
     2161 ctggtttccc tgctgacggt cctgtgatga ccaactcgct gaccgctgcg gactggtgca
     2221 ggtcgaagaa gacttacccc aacgacaaaa ccatcatcag tacctttaag tggagttaca
     2281 ccactggaaa tggcaagcgc taccggagca ctgcgcggac cacctacacc tttgccaagc
     2341 caatggcggc taactatctg aagaaccagc cgatgtacgt gttccgtaag acggagctca
     2401 agcactccaa gaccgagctc aacttcaagg agtggcaaaa ggcctttacc gatgtgatgg
     2461 gcatggacga gctgtacaag taaagatctt tttccctctg ccaaaaatta tggggacatc
     2521 atgaagcccc ttgagcatct gacttctggc taataaagga aatttatttt cattgcaata
     2581 gtgtgttgga attttttgtg tctctcactc ggaaggacat atgggagggc aaatcattta
     2641 aaacatcaga atgagtattt ggtttagagt ttggcaacat atgcccatat gctggctgcc
     2701 atgaacaaag gttggctata aagaggtcat cagtatatga aacagccccc tgctgtccat
     2761 tccttattcc atagaaaagc cttgacttga ggttagattt tttttatatt ttgttttgtg
     2821 ttattttttt ctttaacatc cctaaaattt tccttacatg ttttactagc cagatttttc
     2881 ctcctctcct gactactccc agtcatagct gtccctcttc tcataacttc gtataatgta
     2941 tgctatacga agttatggta ccatggtgag caagggcgag gcagtgatca aggagttcat
     3001 gcggttcaag gtgcacatgg agggctccat gaacggccac gagttcgaga tcgagggcga
     3061 gggcgagggc cgcccctacg agggcaccca gaccgccaag ctgaaggtga ccaagggtgg
     3121 ccccctgccc ttctcctggg acatcctgtc ccctcagttc atgtacggct ccagggcctt
     3181 catcaagcac cccgccgaca tccccgacta ctataagcag tccttccccg agggcttcaa
     3241 gtgggagcgc gtgatgaact tcgaggacgg cggcgccgtg accgtgaccc aggacacctc
     3301 cctggaggac ggcaccctga tctacaaggt gaagctccgc ggcaccaact tccctcctga
     3361 cggccccgta atgcagaaga agacaatggg ctgggaagcg tccaccgagc ggttgtaccc
     3421 cgaggacggc gtgctgaagg gcgacattaa gatggccctg cgcctgaagg acggcggacg
     3481 ctacctggcg gacttcaaga ccacctacaa ggccaagaag cccgtgcaga tgcccggcgc
     3541 ctacaacgtc gaccgcaagt tggacatcac ctcccacaac gaggactaca ccgtggtgga
     3601 acagtacgaa cgctccgagg gccgccactc caccggcggc atggacgagc tgtacaagtt
     3661 cgaaggcagc ggcgctacta acttcagcct gctgaagcag gctggagacg tggaagagaa
     3721 tcctggacct atcgataccg gtcaattggc ggccgcgcat gcgctagcta atttccctct
     3781 gccaaaaatt atggggacat catgaagccc cttgagcatc tgacttctgg ctaataaagg
     3841 aaatttattt tcattgcaat agtgtgttgg aattttttgt gtctctcact cggaaggaca
     3901 tatgggaggg caaatcattt aaaacatcag aatgagtatt tggtttagag tttggcaaca
     3961 tatgcccata tgctggctgc catgaacaaa ggttggctat aaagaggtca tcagtatatg
     4021 aaacagcccc ctgctgtcca ttccttattc catagaaaag ccttgacttg aggttagatt
     4081 ttttttatat tttgttttgt gttatttttt tctttaacat ccctaaaatt ttccttacat
     4141 gttttactag ccagattttt cctcctctcc tgactactcc cagtcatagc tgtccctctt
     4201 ctcttatgga gatccctcga caagcttggc gtaatcatgg tcatagctgt ttcctgtgtg
     4261 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
     4321 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
     4381 ccagtcggga aacctgtcgt gccagcggat ccgcatctca attagtcagc aaccatagtc
     4441 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     4501 catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta
     4561 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctaacttgt
     4621 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     4681 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg
     4741 tctggatccg ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     4801 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     4861 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     4921 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     4981 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     5041 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     5101 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     5161 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     5221 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     5281 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     5341 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     5401 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     5461 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     5521 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     5581 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     5641 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     5701 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     5761 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     5821 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     5881 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     5941 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     6001 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     6061 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     6121 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     6181 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     6241 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     6301 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     6361 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     6421 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     6481 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     6541 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     6601 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     6661 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     6721 ccgaaaagtg ccacctg
//
