LOCUS       sequence-46662-e        4716 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 4716)
  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 4716)
  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: pLSSmOrange-C1
FEATURES             Location/Qualifiers
     source          1..4716
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    8..73
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    197..318
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(234..253)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     288..307
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(325..780)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(462..483)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     674..693
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        807..911
                     /gene="bla"
                     /label=AmpR promoter
     promoter        913..1270
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        913..1104
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(936..956)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      1121..1256
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             1305..2099
                     /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             1305..2099
                     /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(1359..1378)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1969..1988
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(2287..2306)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    2331..2378
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      2707..3295
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3196..3215
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        3470..3773
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3774..3977
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3928..3948
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      4010..4019
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4016..4716,1..7)
                     /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     4429..4448
                     /label=mCherry-F
                     /note="mCherry, forward primer"
ORIGIN
        1 gtacaagtcc ggactcagat ctcgagctca agcttcgaat tctgcagtcg acggtaccgc
       61 gggcccggga tccaccggat ctagataact gatcataatc agccatacca catttgtaga
      121 ggttttactt gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa
      181 tgcaattgtt gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag
      241 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
      301 actcatcaat gtatcttaac gcgtaaattg taagcgttaa tattttgtta aaattcgcgt
      361 taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt
      421 ataaatcaaa agaatagacc gagatagggt tgagtgttgt tccagtttgg aacaagagtc
      481 cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg
      541 gcccactacg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac
      601 taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg
      661 tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag
      721 cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt
      781 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
      841 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
      901 aaaggaagag tcctgaggcg gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg
      961 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     1021 aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     1081 caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc
     1141 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
     1201 ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
     1261 cttttgcaaa gatcgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg
     1321 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac
     1381 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg
     1441 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaagac gaggcagcgc
     1501 ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg
     1561 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc
     1621 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc
     1681 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta
     1741 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg
     1801 cgccagccga actgttcgcc aggctcaagg cgagcatgcc cgacggcgag gatctcgtcg
     1861 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat
     1921 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc
     1981 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta
     2041 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag
     2101 cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt
     2161 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
     2221 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ctagggggag
     2281 gctaactgaa acacggaagg agacaatacc ggaaggaacc cgcgctatga cggcaataaa
     2341 aagacagaat aaaacgcacg gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca
     2401 gggctggcac tctgtcgata ccccaccgag accccattgg ggccaatacg cccgcgtttc
     2461 ttccttttcc ccaccccacc ccccaagttc gggtgaaggc ccagggctcg cagccaacgt
     2521 cggggcggca ggccctgcca tagcctcagg ttactcatat atactttaga ttgatttaaa
     2581 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     2641 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     2701 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     2761 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     2821 tggcttcagc agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca
     2881 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     2941 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     3001 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     3061 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     3121 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     3181 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     3241 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     3301 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     3361 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gccatgcatt agttattaat
     3421 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac
     3481 ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa
     3541 tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
     3601 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc
     3661 ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat
     3721 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc
     3781 ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc
     3841 tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa
     3901 aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg
     3961 tctatataag cagagctggt ttagtgaacc gtcagatccg ctagcggtcg ccaccatggt
     4021 gagcaagggc gaggagaata acatggccat catcaaggag ttcatgcgct tcaaggtgcg
     4081 catggagggc tccgtgaacg gccacgagtt cgagatcgag ggcgagggcg agggccgccc
     4141 ctacgagggc tttcagaccg ttaagctgaa ggtgaccaag ggtggccccc tgcccttcgc
     4201 ctgggacatc ttgtcccctc agttcaccta cggctccaag gcctacgtga agcaccccgc
     4261 cgacatcccc gactacctca agctgtcctt ccccgagggc ttcaagtggg agcgcgtgat
     4321 gaacttcgag gacggcggcg tggtgaccgt gactcaggac tcctccctgc aggacggcga
     4381 gttcatctac aaggtgaagc tgcgcggcac caacttcccc tccgacggcc ccgtaatgca
     4441 gaagaagacc atgggcatgg aggcctcctc cgagcggatg taccccgagg acggcgccct
     4501 gaagggcgag gacaagctca ggctgaagct gaaggacggc ggccactaca cctccgaggt
     4561 caagaccacc tacaaggcca agaagcccgt gcagttgccc ggcgcctaca tcgtcgacat
     4621 caagttggac atcacctccc acaacgagga ctacaccatc gtggaacagt acgaacgcgc
     4681 cgagggccgc cactccaccg gcggcatgga cgagct
//
