LOCUS       sequence-458305-        7276 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transient expression of EGFP FP4-mito in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7276)
  AUTHORS   Scott JA, Shewan AM, den Elzen NR, Loureiro JJ, Gertler FB, Yap AS
  TITLE     Ena/VASP proteins can regulate distinct modes of actin organization 
            at cadherin-adhesive contacts.
  JOURNAL   Mol Biol Cell. 2006 Mar;17(3):1085-95. doi: 10.1091/mbc.e05-07-0644.
            Epub 2005 Dec 21.
  PUBMED    16371509
REFERENCE   2  (bases 1 to 7276)
  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: 
            "10.1091/mbc.e05-07-0644"; journalName: "Mol Biol Cell"; date: 
            "2006-03"; volume: "17"; issue: "3"; pages: "1085-95"
COMMENT     Sequence Label: pcDNA3.1(-)-EGFP-FP4-mito
FEATURES             Location/Qualifiers
     source          1..7276
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      960..969
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(966..968,969..971,972..1682)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(966..968,969..971,972..1682)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1011..1032)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1272..1291)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1619..1640
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2867..2884)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2873..3097
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3143..3571
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3230..3249)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3440..3461
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3580..3600)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3585..3914
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3765..3900
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3827..3846
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3981..4775
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4035..4054)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4645..4664
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4949..5070
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4986..5005)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5040..5059
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5119..5135)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5119..5135)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5132..5154)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5143..5159
                     /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(5167..5173,5174..5191,5192..5197))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5212..5233
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5350..5367)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5521..6109)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5601..5620)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6280..7071,7072..7140))
                     /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     6903..6922
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7141..7245)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaacgg gccctctaga ctcgagcggc cgccactgtg ctggatatct gcagaattcg
      961 ccaccatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     1021 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     1081 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     1141 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
     1201 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     1261 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     1321 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     1381 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
     1441 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     1501 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     1561 accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
     1621 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     1681 agtccggact cagatctatc gtagcttcgg acttcccgcc accacctacg gatgaagagt
     1741 taagacttgc tttgccagag acaccaatgc ttcttggttt taatgctcct gctacatcag
     1801 aaccgagctc attcgaattt ccaccaccac ctacggatga agagttaaga cttgctttgc
     1861 cagagacgcc aatgcttctt ggttttaatg ctcctgctac atcggaaccg agctcgttcg
     1921 aatttccacc gcctccaaca gaagatgaac tagaaatcat ccgggaaaca gcatcctcgc
     1981 tagattctag ttttacaaga ggggatttag ctagtttgag aaatgctatt aatcgccata
     2041 gtcaaaattt ctctgatttc ccaccaatcc caacagaaga agagttgaac gggagaggcg
     2101 gtagaccaac atctgaagaa tttagttcgc tgaatagtgg tgattttaca gatgacgaaa
     2161 acagcgagac aacagaagaa gaaattgatc gcctagctga tttaagagat agaggaacag
     2221 gaaaacactc aagaaatgcg ggttttttac cattaaatcc gtttgctagc agcccggttc
     2281 cttcgttaag tccaaaggta tcgaaaataa gcacaccggc tctgataagt gacataacta
     2341 aaaaaacgcc atttaagaat ccatcacagc cattaaatgt gtttaataaa aaaactacaa
     2401 cgaaaacagt gactaaaaaa ccaacccctg taaagaccgc accaaagcta gcagaacttc
     2461 ctgccacaaa accacaagaa accgtactta gggaaaataa aacacccttt atagaaaaac
     2521 aagcagaaac aaacaagcag tcaattaata tgccgagcct accagtaatc caaaaagaag
     2581 ctacagagag cgataaagag gaaatgaaac cacaaaccga ggaaaaaatg gtagaggaaa
     2641 gcgaatcagc taataacgca aacggaaaaa atcgttctgc tggcattgaa gaaggaaaac
     2701 taattgctaa aagtgcagaa gacgaaaaag cgaaggaaga accagggaac cataggatcg
     2761 taattcttgc aatgttagct attggcgtgt tctctttagg ggcgtttatc aaaattattc
     2821 aattaagaaa aaataattaa aagcttaagt ttaaaccgct gatcagcctc gactgtgcct
     2881 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt
     2941 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg
     3001 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac
     3061 aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc
     3121 tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     3181 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
     3241 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     3301 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag
     3361 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     3421 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
     3481 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     3541 gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt
     3601 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     3661 cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     3721 atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc
     3781 cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg
     3841 ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
     3901 taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct gatcaagaga
     3961 caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg
     4021 cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg
     4081 ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt
     4141 ccggtgccct gaatgaactg caggacgagg cagcgcggct atcgtggctg gccacgacgg
     4201 gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat
     4261 tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat
     4321 ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg
     4381 accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg
     4441 atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc
     4501 tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc
     4561 cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg
     4621 tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg
     4681 gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca
     4741 tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac
     4801 cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga
     4861 aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga
     4921 tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa
     4981 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     5041 tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta
     5101 gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat
     5161 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
     5221 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     5281 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     5341 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     5401 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     5461 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     5521 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     5581 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     5641 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     5701 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     5761 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     5821 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     5881 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     5941 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     6001 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     6061 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     6121 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     6181 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     6241 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     6301 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     6361 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     6421 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     6481 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     6541 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     6601 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     6661 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     6721 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     6781 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     6841 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     6901 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     6961 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     7021 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     7081 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     7141 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     7201 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     7261 agtgccacct gacgtc
//
