LOCUS       sequence-380170-        5696 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human Clathrin Light Chain B tagged with FKBP and mEGFP in
            mammalian cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5696)
  AUTHORS   Obashi K, Sochacki KA, Strub MP, Taraska JW
  TITLE     A conformational switch in clathrin light chain regulates lattice 
            structure and endocytosis at the plasma membrane of mammalian cells.
  JOURNAL   Nat Commun. 2023 Feb 9;14(1):732. doi: 10.1038/s41467-023-36304-7.
  PUBMED    36759616
REFERENCE   2  (bases 1 to 5696)
  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: "2023-02-9"; volume: "14(1):732. doi"; pages: " 
            10.1038/s41467-023-36304-7"
COMMENT     Sequence Label: CLC-FKBP-EGFP
FEATURES             Location/Qualifiers
     source          1..5696
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        68..371
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        372..575
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     526..546
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    598..678
                     /label=MCS
                     /note="multiple cloning site"
     regulatory      680..689
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1337..1339,1340..1660)
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="MGVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNK
                     PFKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVEL
                     LKLE"
     CDS             join(1679..1681,1682..1684,1685..2197)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIE"
     primer_bind     complement(1724..1745)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1985..2004)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2332..2353
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2491..2612
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2528..2547)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2582..2601
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2619..3074)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2756..2777)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2968..2987
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3101..3205
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3207..3564
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3207..3398
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3230..3250)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3415..3550
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3599..4393
                     /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             3599..4393
                     /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(3653..3672)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4263..4282
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4581..4600)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4625..4672
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5001..5589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5490..5509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 catgcattag ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg
       61 agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc
      121 gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt
      181 gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc
      241 atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg
      301 cccagtacat gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg
      361 ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact
      421 cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa
      481 atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta
      541 ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatccgct
      601 agcgctaccg gactcagatc tcgagctcaa gcttcgaatt ctgcagtcga cggtaccgcg
      661 ggcccgggat ccaccggtcg ccaccatggc tgatgacttt ggcttcttct cgtcgtcgga
      721 gagtggtgcc ccggaggcgg cggaggagga cccggcggcc gccttcctgg cccagcagga
      781 gagcgagatt gcaggcatag agaacgacga gggcttcggg gcacctgccg gcagccatgc
      841 ggcccccgca cagccgggcc ccacgagtgg ggctggttct gaggacatgg ggaccacagt
      901 caatggagat gtgtttcagg aggccaacgg tcctgctgat ggctacgcag ccattgccca
      961 ggctgacagg ctgacccagg agcctgagag catccgcaag tggcgagagg agcagaggaa
     1021 acggctgcaa gagctggatg ctgcatctaa ggtcacggaa caggaatggc gggagaaggc
     1081 caagaaggac ctggaggagt ggaaccagcg ccagagtgaa caagtagaga agaacaagat
     1141 caacaaccgg gcatccgagg aggctttcgt gaaggaatcc aaggaggaga ccccaggcac
     1201 agagtgggag aaggtggccc agctatgtga cttcaacccc aagagcagca agcagtgcaa
     1261 agatgtgtcc cgcctgcgct cggtgctcat gtccctgaag cagacgccac tgtcccgcgg
     1321 cggatccgga gggtctatgg gagtgcaggt ggaaaccatc tccccaggag acgggcgcac
     1381 cttccccaag cgcggccaga cctgcgtggt gcactacacc gggatgcttg aagatggaaa
     1441 gaaatttgat tcctcccggg acagaaacaa gccctttaag tttatgctag gcaagcagga
     1501 ggtgatccga ggctgggaag aaggggttgc ccagatgagt gtgggtcaga gagccaaact
     1561 gactatatct ccagattatg cctatggtgc cactgggcac ccaggcatca tcccaccaca
     1621 tgccactctc gtcttcgatg tggagcttct aaaactggaa ggaggatccg gagggtctat
     1681 ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg
     1741 cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg
     1801 caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct
     1861 cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca
     1921 gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt
     1981 caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt
     2041 gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa
     2101 gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg
     2161 catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga
     2221 ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta
     2281 cctgagcacc cagtccaagc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct
     2341 gctggagttc gtgaccgccg ccggctaaag cggccgcgac tctagatcat aatcagccat
     2401 accacatttg tagaggtttt acttgcttta aaaaacctcc cacacctccc cctgaacctg
     2461 aaacataaaa tgaatgcaat tgttgttgtt aacttgttta ttgcagctta taatggttac
     2521 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
     2581 tgtggtttgt ccaaactcat caatgtatct taaggcgtaa attgtaagcg ttaatatttt
     2641 gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat
     2701 cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg ttgttccagt
     2761 ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt
     2821 ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt tggggtcgag
     2881 gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag cttgacgggg
     2941 aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc
     3001 gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc
     3061 gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     3121 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     3181 tcaataatat tgaaaaagga agagtcctga ggcggaaaga accagctgtg gaatgtgtgt
     3241 cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     3301 ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     3361 caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg
     3421 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     3481 tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg aggaggcttt
     3541 tttggaggcc taggcttttg caaagatcga tcaagagaca ggatgaggat cgtttcgcat
     3601 gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg
     3661 ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc
     3721 gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca
     3781 agacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct
     3841 cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga
     3901 tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg
     3961 gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat
     4021 cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga
     4081 gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgagca tgcccgacgg
     4141 cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg
     4201 ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat
     4261 agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct
     4321 cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga
     4381 cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg
     4441 ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt
     4501 ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc
     4561 caccctaggg ggaggctaac tgaaacacgg aaggagacaa taccggaagg aacccgcgct
     4621 atgacggcaa taaaaagaca gaataaaacg cacggtgttg ggtcgtttgt tcataaacgc
     4681 ggggttcggt cccagggctg gcactctgtc gataccccac cgagacccca ttggggccaa
     4741 tacgcccgcg tttcttcctt ttccccaccc caccccccaa gttcgggtga aggcccaggg
     4801 ctcgcagcca acgtcggggc ggcaggccct gccatagcct caggttactc atatatactt
     4861 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     4921 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     4981 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     5041 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     5101 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     5161 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     5221 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     5281 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     5341 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     5401 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     5461 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     5521 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     5581 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     5641 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgc
//
