LOCUS       sequence-325132-        5752 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of zebrafish GFP-tagged Rab40b in mammalian cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5752)
  AUTHORS   Hall TE, Martel N, Ariotti N, Xiong Z, Lo HP, Ferguson C, Rae J, Lim
            YW, Parton RG
  TITLE     In vivo cell biological screening identifies an endocytic capture 
            mechanism for T-tubule formation.
  JOURNAL   Nat Commun. 2020 Jul 24;11(1):3711. doi: 10.1038/s41467-020-17486-w.
  PUBMED    32709891
REFERENCE   2  (bases 1 to 5752)
  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: "Nat 
            Commun."; date: "2020-07-24"; volume: "11(1):3711. doi"; pages: " 
            10.1038/s41467-020-17486-w"
COMMENT     Sequence Label: EGFP-Rab40b in pCSDEST2
FEATURES             Location/Qualifiers
     source          1..5752
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        34..1011
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     promoter        1046..1064
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1046..1063
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     protein_bind    1119..1143
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      1144..1153
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1150..1152,1153..1155,1156..1866)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1150..1152,1153..1155,1156..1866)
                     /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(1195..1216)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1456..1475)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1803..1824
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(1868..1892)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2755..2775)
                     /gene="mutant version of attB"
                     /label=attB3
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     polyA_signal    2815..2949
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2839..2858)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2893..2912
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(3062..3082)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3062..3080)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3101..3117)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3101..3117)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3114..3136)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3125..3141
                     /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(3149..3155,3156..3173,3174..3179))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3194..3215
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3332..3349)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3503..4091)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3583..3602)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4262..5053,5054..5122))
                     /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     4885..4904
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5123..5227)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5253..5708)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5390..5411)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5602..5621
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gtgcgggcct cttcgctatt acgccagtcg accgccaatt caatatggcg tatatggact
       61 catgccaatt caatatggtg gatctggacc tgtgccaatt caatatggcg tatatggact
      121 cgtgccaatt caatatggtg gatctggacc ccagccaatt caatatggcg gacttggcac
      181 catgccaatt caatatggcg gacctggcac tgtgccaact ggggaggggt ctacttggca
      241 cggtgccaag tttgaggagg ggtcttggcc ctgtgccaag tccgccatat tgaattggca
      301 tggtgccaat aatggcggcc atattggcta tatgccagga tcaatatata ggcaatatcc
      361 aatatggccc tatgccaata tggctattgg ccaggttcaa tactatgtat tggccctatg
      421 ccatatagta ttccatatat gggttttcct attgacgtag atagcccctc ccaatgggcg
      481 gtcccatata ccatatatgg ggcttcctaa taccgcccat agccactccc ccattgacgt
      541 caatggtctc tatatatggt ctttcctatt gacgtcatat gggcggtcct attgacgtat
      601 atggcgcctc ccccattgac gtcaattacg gtaaatggcc cgcctggctc aatgcccatt
      661 gacgtcaata ggaccaccca ccattgacgt caatgggatg gctcattgcc cattcatatc
      721 cgttctcacg ccccctattg acgtcaatga cggtaaatgg cccacttggc agtacatcaa
      781 tatctattaa tagtaacttg gcaagtacat tactattgga agtacgccag ggtacattgg
      841 cagtactccc attgacgtca atggcggtaa atggcccgcg atggctgcca agtacatccc
      901 cattgacgtc aatggggagg ggcaatgacg caaatgggcg ttccattgac gtaaatgggc
      961 ggtaggcgtg cctaatggga ggtctatata agcaatgctc gtttagggaa ccgccattct
     1021 gcctggggac gtcggagcaa gcttgattta ggtgacacta tagaatacaa gctacttgtt
     1081 ctttttgcag gatcccatcg attcgaattc aaggatcaac aagtttgtac aaaaaagcag
     1141 gctgccacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc
     1201 gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat
     1261 gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc
     1321 tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac
     1381 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc
     1441 accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc
     1501 gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc
     1561 ctggggcaca agctggagta caactacaac agccacaacg tctatatcat ggccgacaag
     1621 cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg
     1681 cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc
     1741 gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat
     1801 cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg
     1861 tacaagcacc cagctttctt gtacaaagtg gtgggtcatc gggctgatgt gaagatgaac
     1921 cacaggagca gcccagcaaa agcttatgat tttcttctga aattcctact ggttggagac
     1981 agtgatgtgg ggaaagggga aatactggcc agtcttcagg atgggtccag tgagtcaccc
     2041 tatggataca acatgggaat cgactacaag actactacca ttctcctgga tggacgcagg
     2101 gttaagcttc aactgtggga cacctcagga caaggcagat tctgtaccat attccgctcc
     2161 tattccagag gagcacaggg agtcatcctt gtttatgaca tcacaaaccg gtggtctttc
     2221 gacggcatcg acagatggat taaagagatt gatgagcatg caccaggtgt ccccaagatc
     2281 ctggtgggca acaggcttca tctggcctat aagcgtcagg tgaccaccga acatgcgcag
     2341 gcctttgcag agcgattggg cgtcacgttt tttgaggtca gccctctctg taacttcaac
     2401 atcaccgagt ccttcactga actggcccgc attgtgctca tgaggcacgg catggagcga
     2461 ctctggagac ccaataaagt gttgagtctg caggacctgt gctgccgttc tatcgtttcc
     2521 tgcactccag tgcatctcgt ggacaaactt ccccttcctg ttgccttaaa gagtcatctg
     2581 aagtcctttt ccatggctaa cggcctggtc aacaccagga tgatgcacgg ccgttcctat
     2641 tcggtcatac ctagcagtgc taaaaaacgg aacggaacca aaaaatccaa aatcatctat
     2701 cctccactga gcccatccca gcactgcgca cggacaagct gcaagatttc atagcaactt
     2761 tattatacat agttgcactc gagcctctag aactatagtg agtcgtatta cgtagatcca
     2821 gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa
     2881 tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat
     2941 aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggaggtgtgg
     3001 gaggtttttt aattcgcggc cgcggcgcca atgcattggg cccggtaccc agcttttgtt
     3061 ccctttagtg agggttaatt gcgcgcttgg cgtaatcatg gtcatagctg tttcctgtgt
     3121 gaaattgtta tccgctcaca attccacaca acatacgagc cgggagcata aagtgtaaag
     3181 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     3241 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     3301 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     3361 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     3421 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     3481 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     3541 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     3601 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     3661 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     3721 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     3781 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     3841 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     3901 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     3961 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     4021 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     4081 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     4141 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     4201 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     4261 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     4321 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     4381 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     4441 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     4501 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     4561 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     4621 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     4681 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     4741 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     4801 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     4861 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     4921 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     4981 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     5041 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     5101 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     5161 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     5221 ttccgcgcac atttccccga aaagtgccac ctaaattgta agcgttaata ttttgttaaa
     5281 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
     5341 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
     5401 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
     5461 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
     5521 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
     5581 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
     5641 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
     5701 gggcgcgtcc cattcgccat tcaggctgcg caactgttgg gaagggcgat cg
//
