LOCUS       sequence-46701-e        5094 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5094)
  AUTHORS   Shcherbakova DM, Hink MA, Joosen L, Gadella TW, Verkhusha VV
  TITLE     An Orange Fluorescent Protein with a Large Stokes Shift for 
            Single-Excitation Multicolor FCCS and FRET Imaging.
  JOURNAL   J Am Chem Soc. 2012 May 9;134(18):7913-23. doi: 10.1021/ja3018972
  PUBMED    22486524
REFERENCE   2  (bases 1 to 5094)
  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"; doi: "10"; journalName: 
            "J Am Chem Soc"; date: "2012-05-9- 9"; volume: "134"; issue: "18"; 
            pages: "7913-23"
COMMENT     Sequence Label: pH2B-LSSmOrange
FEATURES             Location/Qualifiers
     source          1..5094
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1044..1053
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1050..1760
                     /codon_start=1
                     /product="monomeric orange fluorescent protein with a large
                     Stokes shift (Shcherbakova et al., 2012)"
                     /label=LSSmOrange
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEENNMAIIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEG
                     FQTVKLKVTKGGPLPFAWDILSPQFTYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGMEASSERMYPEDGALK
                     GEDKLRLKLKDGGHYTSEVKTTYKAKKPVQLPGAYIVDIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     1463..1482
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    1882..2003
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1919..1938)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1973..1992
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2010..2465)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2147..2168)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2359..2378
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2492..2596
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2598..2955
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2598..2789
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2621..2641)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2806..2941
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2990..3784
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2990..3784
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3044..3063)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3654..3673
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3972..3991)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4016..4063
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4392..4980
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4881..4900
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgagct caagcttcga attctgcagt cgacggtacc gccaccatgc
      661 cagagccagc gaagtctgct cccgccccga aaaagggctc caagaaggcg gtgactaagg
      721 cgcagaagaa agacggcaag agcgcagcgc accgcaagga gagctattcc atctatgtgt
      781 acaaggttct gaagcaggtc caccctgaca ccggcatttc gtccaaggcc atgggcatca
      841 tgaattcgtt tgtgaacgac attttcgagc gcatcgcagg tgaggcttcc cgccttgcgc
      901 attacaacaa gcgctcgacc atcacctcca gggagatcca gacggccgtg cgcctgctgc
      961 tgcctgggga gttggccaag cacgccgtgt ccgagggtac taaggccgtc accaagtaca
     1021 ccagcgctaa ggatccaccg gtcgccacca tggtgagcaa gggcgaggag aataacatgg
     1081 ccatcatcaa ggagttcatg cgcttcaagg tgcgcatgga gggctccgtg aacggccacg
     1141 agttcgagat cgagggcgag ggcgagggcc gcccctacga gggctttcag accgttaagc
     1201 tgaaggtgac caagggtggc cccctgccct tcgcctggga catcttgtcc cctcagttca
     1261 cctacggctc caaggcctac gtgaagcacc ccgccgacat ccccgactac ctcaagctgt
     1321 ccttccccga gggcttcaag tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga
     1381 ccgtgactca ggactcctcc ctgcaggacg gcgagttcat ctacaaggtg aagctgcgcg
     1441 gcaccaactt cccctccgac ggccccgtaa tgcagaagaa gaccatgggc atggaggcct
     1501 cctccgagcg gatgtacccc gaggacggcg ccctgaaggg cgaggacaag ctcaggctga
     1561 agctgaagga cggcggccac tacacctccg aggtcaagac cacctacaag gccaagaagc
     1621 ccgtgcagtt gcccggcgcc tacatcgtcg acatcaagtt ggacatcacc tcccacaacg
     1681 aggactacac catcgtggaa cagtacgaac gcgccgaggg ccgccactcc accggcggca
     1741 tggacgagct gtacaagtaa gcggccgcga ctctagatca taatcagcca taccacattt
     1801 gtagaggttt tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa
     1861 atgaatgcaa ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc
     1921 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     1981 tccaaactca tcaatgtatc ttaaggcgta aattgtaagc gttaatattt tgttaaaatt
     2041 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat
     2101 cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa
     2161 gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg
     2221 cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa
     2281 agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc
     2341 gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag
     2401 tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg
     2461 cgcgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     2521 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     2581 ttgaaaaagg aagagtcctg aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg
     2641 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     2701 tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     2761 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     2821 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     2881 gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     2941 ctaggctttt gcaaagatcg atcaagagac aggatgagga tcgtttcgca tgattgaaca
     3001 agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg
     3061 ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg
     3121 cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aagacgaggc
     3181 agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt
     3241 cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc
     3301 atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca
     3361 tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc
     3421 acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg
     3481 gctcgcgcca gccgaactgt tcgccaggct caaggcgagc atgcccgacg gcgaggatct
     3541 cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc
     3601 tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc
     3661 tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta
     3721 cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt
     3781 ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga
     3841 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac
     3901 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccctagg
     3961 gggaggctaa ctgaaacacg gaaggagaca ataccggaag gaacccgcgc tatgacggca
     4021 ataaaaagac agaataaaac gcacggtgtt gggtcgtttg ttcataaacg cggggttcgg
     4081 tcccagggct ggcactctgt cgatacccca ccgagacccc attggggcca atacgcccgc
     4141 gtttcttcct tttccccacc ccacccccca agttcgggtg aaggcccagg gctcgcagcc
     4201 aacgtcgggg cggcaggccc tgccatagcc tcaggttact catatatact ttagattgat
     4261 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     4321 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     4381 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     4441 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     4501 gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta
     4561 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     4621 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     4681 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     4741 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     4801 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     4861 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     4921 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     4981 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     5041 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgccat gcat
//
