LOCUS       sequence-388861-        8129 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express eGFP-RAPH1 lacking amino acids 536-587 in mammalian cells. 
            This constructs does not localise to filopodia anymore..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8129)
  AUTHORS   Popovic A, Miihkinen M, Ghimire S, Saup R, Gronloh MLB, Ball NJ, 
            Goult BT, Ivaska J, Jacquemet G
  TITLE     Myosin-X recruits lamellipodin to filopodia tips.
  JOURNAL   J Cell Sci. 2023 Mar 1;136(5):jcs260574. doi: 10.1242/jcs.260574. 
            Epub 2023 Mar 2.
  PUBMED    36861887
REFERENCE   2  (bases 1 to 8129)
  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: "J Cell 
            Sci."; date: "2023-03-1"; volume: "136(5):jcs260574. doi"; pages: " 
            10.1242/jcs.260574. Epub 2023 Mar 2"
COMMENT     Sequence Label: GFP-RAPH1-Delta-536-587
FEATURES             Location/Qualifiers
     source          1..8129
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        172..375
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     326..346
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      414..423
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(420..422,423..425,426..1136)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(420..422,423..425,426..1136)
                     /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(465..486)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(726..745)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1073..1094
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      4590..4599
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    4917..5038
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4954..4973)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5008..5027
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5045..5500)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5182..5203)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5394..5413
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5527..5631
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5633..5990
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5633..5824
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5656..5676)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5841..5976
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6025..6819
                     /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             6025..6819
                     /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(6079..6098)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6689..6708
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(7007..7026)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    7051..7098
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      7427..8015
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7916..7935
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg
      121 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg
      181 ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc
      241 caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa
      301 tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc
      361 tatataagca gagctggttt agtgaaccgt cagatccgct agcgctaccg gtcgccacca
      421 tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg
      481 gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg
      541 gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc
      601 tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc
      661 agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct
      721 tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg
      781 tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca
      841 agctggagta caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg
      901 gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg
      961 accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact
     1021 acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc
     1081 tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtccg
     1141 gactcagatc tcgagccgcg atggagcagc tatcagatga agaaattgat catggtgctg
     1201 aagaagacag tgacaaggaa gatcaggacc tggacaaaat gtttggagcc tggcttggag
     1261 aactagacaa actcactcag agtttggatt ctgacaagcc catggaacca gtaaaaagat
     1321 ctcctcttcg ccaggaaaca aacatggcca acttttctta ccgcttctcc atatacaact
     1381 tgaatgaagc tctgaatcag ggagagactg tggatctgga tgccttgatg gctgatcttt
     1441 gctctataga gcaggagctc agcagcattg gttcaggaaa cagtaagcgt caaatcacag
     1501 aaacgaaagc tactcagaaa ttgcctgtta gccgacatac attgaaacat ggcaccttga
     1561 aaggattatc ttcttcatct aataggatag ctaaaccttc ccatgccagc tactccttgg
     1621 acgacgtcac tgcacagtta gaacaggcct ctttgagtat ggatgaggct gctcagcaat
     1681 ctgtactaga agatactaaa cccttagtaa ctaatcagca cagaagaacc gcgtcagcag
     1741 gcacagtgag tgatgctgaa gtacactcta ttagtaattc ctcccattcc agcatcactt
     1801 ccgcagcctc cagcatggac tctttggata ttgataaagt aacacgccct caagagctgg
     1861 atttgacaca tcaagggcag ccaattactg aggaagaaca ggcagcaaaa ttgaaagctg
     1921 agaagatcag agttgcccta gagaaaatta aagaggcaca agtgaaaaag ctggtgatca
     1981 gagtccacat gtctgatgac agttctaaaa caatgatggt ggatgagagg cagacagtaa
     2041 gacaagtact ggataacctg atggacaaat cccactgcgg ttatagttta gactggtcac
     2101 tggtagaaac cgtttctgaa ttacaaatgg agagaatctt tgaagaccat gaaaacttgg
     2161 ttgaaaatct tcttaattgg acaagagata gccaaaacaa gcttatattt atggagcgta
     2221 tagaaaaata tgcacttttc aaaaacccac agaattatct tttggggaaa aaggaaacag
     2281 ctgagatggc agatagaaac aaagaagtcc tcttggagga atgtttttgt ggaagttctg
     2341 taactgtacc agaaattgaa ggagtccttt ggttgaagga tgatggcaag aagtcctgga
     2401 aaaagcgtta ttttctcttg cgagcatctg gtatctacta tgttcccaaa ggaaaagcaa
     2461 aggtctctcg ggatctggtg tgctttctcc agctggatca tgtcaacgtt tattatggcc
     2521 aggactatcg gaacaaatac aaagcaccta cagactattg tctggtgctg aagcatccac
     2581 aaatccagaa gaaatctcaa tatatcaaat acctttgttg tgatgatgtg aggacactgc
     2641 atcagtgggt caatgggatc cgcattgcaa agtatgggaa gcagctctat atgaactacc
     2701 aagaagcctt gaagaggaca gagtcagcct atgattggac ttccttatcc agctccagca
     2761 ttaaatcgga agagtccagc aaggccagaa tggagtctat gaatcggccc tacacttcac
     2821 ttgtgccccc tttatccccg caacctaaga tagtcacccc ctacactgct tcacagcctt
     2881 caccacctct acctcctccg ccacccccac ctcctcctcc accaccccct ccaccacccc
     2941 ctcctccccc actccccagc cagtctgcac cttctgcagg ctcagcagcc ccaatgttcg
     3001 tcaagtacag cacaataaca cggctacaga atgcgtctca gcattcaggg gccctgttta
     3061 agccgccaac acccccagtg atgcagtcac agtcagtgaa gcctcagatc ctggtacccc
     3121 ccaatggagt tgttccacca ccccctcccc ctcctccacc cccaacccca ggctctgcca
     3181 tggcccagct aaagcctgca ccgtgtgccc catcccttcc acagttcagt gccccgcctc
     3241 ctccactgaa gatccatcaa gttcagcata ttactcaggt ggctccccca acaccccccc
     3301 cacctcctcc tatccctgca cccctccctc cccaagctcc cccaaaaccc cttgtgacca
     3361 tccccgcacc aaccagcacc aagactgtgg cacctgttgt gactcaagct gcaccaccca
     3421 cacctactcc tccagtgccc ccagcaaaaa agcagccagc tttccctgct tcttacattc
     3481 caccctctcc ccctacccct cctgttccag tacccccgcc aacattaccc aagcaacaga
     3541 gcttctgtgc aaaaccccct ccctctccac tgtcaccggt gccctcggtc gtgaagcaga
     3601 tagccagcca gtttccaccc cctccaactc cccctgccat ggaatctcag cccttaaagc
     3661 ctgtcccagc aaatgtagct ccacagtccc ctcctgcagt aaaagcaaag cccaagtggc
     3721 agcccagctc catcccagtc ccttctccgg acttccctcc tccccctcct gaaagcagcc
     3781 tggtgtttcc tcctccaccc ccatcacctg tcccagcccc accaccgcca cctccaccca
     3841 cagcttctcc tacccctgac aaaagtggat ctccaggcaa aaagaccagt aagacgtcca
     3901 gccctggggg aaagaaacca cccccaaccc cacagcgcaa ctccagcatt aaatccagca
     3961 gtggtgcaga gcaccccgag cccaagagac cctcggtgga cagtctagtc agcaagttta
     4021 caccgccagc agaatcaggg tctcccagca aggagaccct accacctcct gcagcacccc
     4081 ccaagcctgg aaaactcaat ctttctggag tcaaccttcc tggagttctc caacaagggt
     4141 gtgtgtcagc aaaagcccct gttctgagtg ggcgtggaaa ggactccgtg gtggaatttc
     4201 cttctcctcc atccgattct gattttccac cccctccacc tgaaacagac cttcctctgc
     4261 cccccattga gattccagca gttttctcgg gaaacacctc tccaaaagtg gcagtcgtta
     4321 atcctcaacc acaacaatgg tctaaaatgt cagtgaagaa ggcccctcca cccacacgac
     4381 ccaaacggaa tgatagcacc cgcctcactc aagctgagat ttctgagcag ccaacaatgg
     4441 ccacagttgt gccacaagtg cccacctctc ccaaatccag ccttagtgtc cagcctggat
     4501 tcctggctga cctcaacagg acactgcaac gaaagtccat cactcggcac ggctcactct
     4561 cctcccgcat gtccagagca gaaccaacag ccaccatgga tgatatggca ttgcctccac
     4621 caccccctga actgctgtct gatcaacaga aggctggtta cggaggcagt catatatcag
     4681 gctatgcaac gttgcggaga ggaccccctc ctgctccccc caaaagagac cagaacacca
     4741 agctctccag agactggtag ccaccatagg actttaccgc gggcccggga tccaccggat
     4801 ctagataact gatcataatc agccatacca catttgtaga ggttttactt gctttaaaaa
     4861 acctcccaca cctccccctg aacctgaaac ataaaatgaa tgcaattgtt gttgttaact
     4921 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     4981 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttaac
     5041 gcgtaaattg taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc
     5101 tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc
     5161 gagatagggt tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac
     5221 tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca
     5281 ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg
     5341 agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag
     5401 aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc
     5461 accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt caggtggcac ttttcgggga
     5521 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     5581 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tcctgaggcg
     5641 gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag
     5701 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc
     5761 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag
     5821 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
     5881 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc
     5941 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa gatcgatcaa
     6001 gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg
     6061 gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat cggctgctct
     6121 gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac
     6181 ctgtccggtg ccctgaatga actgcaagac gaggcagcgc ggctatcgtg gctggccacg
     6241 acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg
     6301 ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa
     6361 gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca
     6421 ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt
     6481 gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc
     6541 aggctcaagg cgagcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc
     6601 ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg
     6661 ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt
     6721 ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag
     6781 cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa
     6841 tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct
     6901 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
     6961 gggatctcat gctggagttc ttcgcccacc ctagggggag gctaactgaa acacggaagg
     7021 agacaatacc ggaaggaacc cgcgctatga cggcaataaa aagacagaat aaaacgcacg
     7081 gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca gggctggcac tctgtcgata
     7141 ccccaccgag accccattgg ggccaatacg cccgcgtttc ttccttttcc ccaccccacc
     7201 ccccaagttc gggtgaaggc ccagggctcg cagccaacgt cggggcggca ggccctgcca
     7261 tagcctcagg ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     7321 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     7381 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     7441 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     7501 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     7561 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     7621 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     7681 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     7741 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     7801 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     7861 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     7921 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     7981 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     8041 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     8101 ctgtggataa ccgtattacc gccatgcat
//
