LOCUS       sequence-181643-        5414 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of a fluorescent protein, monomeric Apple 
            (mApple). Note: Addgene identified several mutations compared to the
            final published sequence..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5414)
  AUTHORS   Shaner NC, Lin MZ, McKeown MR, Steinbach PA, Hazelwood KL, Davidson 
            MW, Tsien RY
  TITLE     Improving the photostability of bright monomeric orange and red 
            fluorescent proteins.
  JOURNAL   Nat Methods. 2008 Jun;5(6):545-51. doi: 10.1038/nmeth.1209. Epub 
            2008 May 4.
  PUBMED    18454154
REFERENCE   2  (bases 1 to 5414)
  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: 
            "10.1038/nmeth.1209"; journalName: "Nat Methods"; date: "2008-06"; 
            volume: "5"; issue: "6"; pages: "545-51"
COMMENT     Sequence Label: pBAD mApple
FEATURES             Location/Qualifiers
     source          1..5414
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             18..35
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             39..71
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             join(75..83,84..98)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             105..815
                     /codon_start=1
                     /product="photostable monomeric derivative of DsRed (Shaner
                     et al., 2008)"
                     /label=mApple
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEENNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEA
                     FQTAKLKVTKGGPLPFAWDILSPQFMYGSKVYIKHPADIPDYFKLSFPEGFKWERVMNF
                     EDGGIIHVNQDSSLQDGVFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     SEIKKRLKLKDGGHYASEVKTTYKAKKPVQLPGAYIVDIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     518..537
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(861..878)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1031..1117
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1209..1236
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1255..1346
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1347..1415,1416..2207)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1565..1584)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2249..2704
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2336..2355)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2546..2567
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     rep_origin      2815..3403
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3304..3323
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3557..3574
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3589..3729
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3655..3674)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3815..3837
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3975..3994
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     CDS             complement(4174..5052)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        5079..5363
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     5286..5305
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     RBS             5390..5412
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
ORIGIN
        1 tacatatgcg gggttctcat catcatcatc atcatggtat ggctagcatg actggtggac
       61 agcaaatggg tcgggatctg tacgacgatg acgataagga tccgatggtg agcaagggcg
      121 aggagaataa catggccatc atcaaggagt tcatgcgctt caaggtgcac atggagggct
      181 ccgtgaacgg ccacgagttc gagatcgagg gcgagggcga gggccgcccc tacgaggcct
      241 ttcagaccgc taagctgaag gtgaccaagg gtggccccct gcccttcgcc tgggacatcc
      301 tgtcccctca gttcatgtac ggctccaagg tctacattaa gcacccagcc gacatccccg
      361 actacttcaa gctgtccttc cccgagggct tcaagtggga gcgcgtgatg aacttcgagg
      421 acggcggcat tattcacgtt aaccaggact cctccctgca ggacggcgtg ttcatctaca
      481 aggtgaagct gcgcggcacc aacttcccct ccgacggccc cgtaatgcag aagaagacca
      541 tgggctggga ggcctcctcc gagcggatgt accccgagga cggcgccctg aagagcgaga
      601 tcaagaagag gctgaagctg aaggacggcg gccactacgc ctccgaggtc aagaccacct
      661 acaaggccaa gaagcccgtg cagctgcccg gcgcctacat cgtcgacatc aagttggaca
      721 tcacctccca caacgaggac tacaccatcg tggaacagta cgaacgcgcc gagggccgcc
      781 actccaccgg cggcatggac gagctgtaca agtaagaatt cgaagcttgg ctgttttggc
      841 ggatgagaga agattttcag cctgatacag attaaatcag aacgcagaag cggtctgata
      901 aaacagaatt tgcctggcgg cagtagcgcg gtggtcccac ctgaccccat gccgaactca
      961 gaagtgaaac gccgtagcgc cgatggtagt gtggggtctc cccatgcgag agtagggaac
     1021 tgccaggcat caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg
     1081 ttgtttgtcg gtgaacgctc tcctgagtag gacaaatccg ccgggagcgg atttgaacgt
     1141 tgcgaagcaa cggcccggag ggtggcgggc aggacgcccg ccataaactg ccaggcatca
     1201 aattaagcag aaggccatcc tgacggatgg cctttttgcg tttctacaaa ctcttttgtt
     1261 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     1321 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     1381 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     1441 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     1501 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     1561 ttctgctatg tggcgcggta ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc
     1621 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     1681 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     1741 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     1801 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     1861 caaacgacga gcgtgacacc acgatgcctg cagcaatggc aacaacgttg cgcaaactat
     1921 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     1981 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     2041 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     2101 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     2161 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     2221 tttactcata tatactttag attgatttac gcgccctgta gcggcgcatt aagcgcggcg
     2281 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct
     2341 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
     2401 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
     2461 gatttgggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg
     2521 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac
     2581 cctatctcgg gctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta
     2641 aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgtttaca
     2701 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     2761 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     2821 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     2881 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     2941 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa
     3001 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     3061 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     3121 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     3181 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     3241 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     3301 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     3361 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     3421 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     3481 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     3541 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta
     3601 ttttctcctt acgcatctgt gcggtatttc acaccgcatc tggtgcactc tcagtacaat
     3661 ctgctctgat gccgcatagt taagccagta tacactccgc tatcgctacg tgactgggtc
     3721 atggctgcgc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     3781 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     3841 tcaccgtcat caccgaaacg cgcgaggcag aaggagatgg cgcccaacag tcccccggcc
     3901 acggggcctg ccaccatacc cacgccgaaa caagcgctca tgagcccgaa gtggcgagcc
     3961 cgatcttccc catcggtgat gtcggcgata taggcgccag caaccgcacc tgtggcgccg
     4021 gtgatgccgg ccacgatgcg tccggcgtag aggatctgct catgtttgac agcttatcat
     4081 cgatgcataa tgtgcctgtc aaatggacga agcagggatt ctgcaaaccc tatgctactc
     4141 cgtcaagccg tcaattgtct gattcgttac caattatgac aacttgacgg ctacatcatt
     4201 cactttttct tcacaaccgg cacggaactc gctcgggctg gccccggtgc attttttaaa
     4261 tacccgcgag aaatagagtt gatcgtcaaa accaacattg cgaccgacgg tggcgatagg
     4321 catccgggtg gtgctcaaaa gcagcttcgc ctggctgata cgttggtcct cgcgccagct
     4381 taagacgcta atccctaact gctggcggaa aagatgtgac agacgcgacg gcgacaagca
     4441 aacatgctgt gcgacgctgg cgatatcaaa attgctgtct gccaggtgat cgctgatgta
     4501 ctgacaagcc tcgcgtaccc gattatccat cggtggatgg agcgactcgt taatcgcttc
     4561 catgcgccgc agtaacaatt gctcaagcag atttatcgcc agcagctccg aatagcgccc
     4621 ttccccttgc ccggcgttaa tgatttgccc aaacaggtcg ctgaaatgcg gctggtgcgc
     4681 ttcatccggg cgaaagaacc ccgtattggc aaatattgac ggccagttaa gccattcatg
     4741 ccagtaggcg cgcggacgaa agtaaaccca ctggtgatac cattcgcgag cctccggatg
     4801 acgaccgtag tgatgaatct ctcctggcgg gaacagcaaa atatcacccg gtcggcaaac
     4861 aaattctcgt ccctgatttt tcaccacccc ctgaccgcga atggtgagat tgagaatata
     4921 acctttcatt cccagcggtc ggtcgataaa aaaatcgaga taaccgttgg cctcaatcgg
     4981 cgttaaaccc gccaccagat gggcattaaa cgagtatccc ggcagcaggg gatcattttg
     5041 cgcttcagcc atacttttca tactcccgcc attcagagaa gaaaccaatt gtccatattg
     5101 catcagacat tgccgtcact gcgtctttta ctggctcttc tcgctaacca aaccggtaac
     5161 cccgcttatt aaaagcattc tgtaacaaag cgggaccaaa gccatgacaa aaacgcgtaa
     5221 caaaagtgtc tataatcacg gcagaaaagt ccacattgat tatttgcacg gcgtcacact
     5281 ttgctatgcc atagcatttt tatccataag attagcggat actacctgac gctttttatc
     5341 gcaactctct actgtttctc catacccgtt tttttgggct agaaataatt ttgtttaact
     5401 ttaagaagga gata
//
