LOCUS       sequence-380179-        5835 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human Clathrin Light Chain B tagged with SNAP-tag and 
            mEGFP in mammalian cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5835)
  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 5835)
  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: EGFP-CLC-SNAP
FEATURES             Location/Qualifiers
     source          1..5835
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        100..291
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     248..268
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        complement(293..397)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      424..879
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(511..530)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     721..742
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    886..1007
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(897..916)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     951..970
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(1119..1664)
                     /codon_start=1
                     /product="human O6-alkylguanine-DNA-alkyltransferase 
                     variant that forms covalent bonds with benzylguanine 
                     derivatives"
                     /label=SNAP-tag(R)
                     /note="optimized for mammalian expression"
                     /translation="MDKDCEMKRTTLDSPLGKLELSGCEQGLHEIKLLGKGTSAADAVE
                     VPAPAAVLGGPEPLMQATAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLK
                     VVKFGEVISYQQLAALAGNPAATAAVKTALSGNPVPILIPCHRVVSSSGAVGGYEGGLA
                     VKEWLLAHEGHRLGKPGLG"
     primer_bind     complement(2343..2364)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             complement(join(2499..3011,3012..3014,3015..3017))
                     /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     2692..2711
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2951..2972
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      3014..3023
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(3062..3265)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(3091..3111)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        3266..3569
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(3744..4332)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3824..3843)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    4661..4708
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     4733..4752
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(4940..5734)
                     /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             complement(4940..5734)
                     /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(5051..5070)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5661..5680
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(join(5769..5835,1..291))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(5783..5835,1..83)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 gcctcggcct ctgcataaat aaaaaaaatt agtcagccat ggggcggaga atgggcggaa
       61 ctgggcggag ttaggggcgg gatgggcgga gttaggggcg ggactatggt tgctgactaa
      121 ttgagatgca tgctttgcat acttctgcct gctggggagc ctggggactt tccacacctg
      181 gttgctgact aattgagatg catgctttgc atacttctgc ctgctgggga gcctggggac
      241 tttccacacc ctaactgaca cacattccac agctggttct ttccgcctca ggactcttcc
      301 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
      361 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
      421 ctgacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
      481 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
      541 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
      601 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
      661 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
      721 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
      781 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
      841 ttaacgcgaa ttttaacaaa atattaacgc ttacaattta cgcgttaaga tacattgatg
      901 agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg
      961 atgctattgc tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt
     1021 gcattcattt tatgtttcag gttcaggggg aggtgtggga ggttttttaa agcaagtaaa
     1081 acctctacaa atgtggtatg gctgattatg atcagttaac ccagcccagg cttgcccagt
     1141 ctgtggccct cgtgggccag cagccactct ttcacggcga gcccgccctc gtagcccccc
     1201 acggcgccag agctagacac cacccggtgg caggggatca gaatgggcac gggatttccg
     1261 ctcagggcgg ttttcacggc ggcggtggcg gcgggattgc cggccagggc ggccagctgc
     1321 tggtagctga tgacctctcc gaacttcacc actttcagca gtttccacag cacctggcgg
     1381 gtaaagctct cctgctggaa cactgggtgg tgcagggctg gcacagggaa ctcctcgatg
     1441 gcctcaggct ggtgaaagta ggcgttgagc caggcggtgg cctgcatcag tggctctggt
     1501 ccgcccagca cggcggctgg ggcaggcact tccacggcgt cggcggcaga tgttcctttg
     1561 cccagcagct tgatctcgtg caggccctgt tcgcacccag acagttccag cttgcccaga
     1621 gggctatcca gggtggtgcg cttcatttcg cagtctttgt ccatagaccc tccggatccg
     1681 ccgcgggaca gtggcgtctg cttcagggac atgagcaccg agcgcaggcg ggacacatct
     1741 ttgcactgct tgctgctctt ggggttgaag tcacatagct gggccacctt ctcccactct
     1801 gtgcctgggg tctcctcctt ggattccttc acgaaagcct cctcggatgc ccggttgttg
     1861 atcttgttct tctctacttg ttcactctgg cgctggttcc actcctccag gtccttcttg
     1921 gccttctccc gccattcctg ttccgtgacc ttagatgcag catccagctc ttgcagccgt
     1981 ttcctctgct cctctcgcca cttgcggatg ctctcaggct cctgggtcag cctgtcagcc
     2041 tgggcaatgg ctgcgtagcc atcagcagga ccgttggcct cctgaaacac atctccattg
     2101 actgtggtcc ccatgtcctc agaaccagcc ccactcgtgg ggcccggctg tgcgggggcc
     2161 gcatggctgc cggcaggtgc cccgaagccc tcgtcgttct ctatgcctgc aatctcgctc
     2221 tcctgctggg ccaggaaggc ggccgccggg tcctcctccg ccgcctccgg ggcaccactc
     2281 tccgacgacg agaagaagcc aaagtcatca gccatagacc ctccggatcc gccggcggcg
     2341 gtcacgaact ccagcaggac catgtgatcg cgcttctcgt tggggtcttt gctcagcttg
     2401 gactgggtgc tcaggtagtg gttgtcgggc agcagcacgg ggccgtcgcc gatgggggtg
     2461 ttctgctggt agtggtcggc gagctgcacg ctgccgtcct cgatgttgtg gcggatcttg
     2521 aagttcacct tgatgccgtt cttctgcttg tcggccatga tatagacgtt gtggctgttg
     2581 tagttgtact ccagcttgtg ccccaggatg ttgccgtcct ccttgaagtc gatgcccttc
     2641 agctcgatgc ggttcaccag ggtgtcgccc tcgaacttca cctcggcgcg ggtcttgtag
     2701 ttgccgtcgt ccttgaagaa gatggtgcgc tcctggacgt agccttcggg catggcggac
     2761 ttgaagaagt cgtgctgctt catgtggtcg gggtagcggc tgaagcactg cacgccgtag
     2821 gtcagggtgg tcacgagggt gggccagggc acgggcagct tgccggtggt gcagatgaac
     2881 ttcagggtca gcttgccgta ggtggcatcg ccctcgccct cgccggacac gctgaacttg
     2941 tggccgttta cgtcgccgtc cagctcgacc aggatgggca ccaccccggt gaacagctcc
     3001 tcgcccttgc tcaccatggt ggcgaccggt agcgctagcg gatctgacgg ttcactaaac
     3061 cagctctgct tatatagacc tcccaccgta cacgcctacc gcccatttgc gtcaatgggg
     3121 cggagttgtt acgacatttt ggaaagtccc gttgattttg gtgccaaaac aaactcccat
     3181 tgacgtcaat ggggtggaga cttggaaatc cccgtgagtc aaaccgctat ccacgcccat
     3241 tgatgtactg ccaaaaccgc atcaccatgg taatagcgat gactaatacg tagatgtact
     3301 gccaagtagg aaagtcccat aaggtcatgt actgggcata atgccaggcg ggccatttac
     3361 cgtcattgac gtcaataggg ggcgtacttg gcatatgata cacttgatgt actgccaagt
     3421 gggcagttta ccgtaaatac tccacccatt gacgtcaatg gaaagtccct attggcgtta
     3481 ctatgggaac atacgtcatt attgacgtca atgggcgggg gtcgttgggc ggtcagccag
     3541 gcgggccatt taccgtaagt tatgtaacgc ggaactccat atatgggcta tgaactaatg
     3601 accccgtaat tgattactat taataactaa tgcatggcgg taatacggtt atccacagaa
     3661 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     3721 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     3781 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     3841 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     3901 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     3961 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     4021 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     4081 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     4141 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     4201 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     4261 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     4321 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     4381 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     4441 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaacc tgaggctatg
     4501 gcagggcctg ccgccccgac gttggctgcg agccctgggc cttcacccga acttgggggg
     4561 tggggtgggg aaaaggaaga aacgcgggcg tattggcccc aatggggtct cggtggggta
     4621 tcgacagagt gccagccctg ggaccgaacc ccgcgtttat gaacaaacga cccaacaccg
     4681 tgcgttttat tctgtctttt tattgccgtc atagcgcggg ttccttccgg tattgtctcc
     4741 ttccgtgttt cagttagcct ccccctaggg tgggcgaaga actccagcat gagatccccg
     4801 cgctggagga tcatccagcc ggcgtcccgg aaaacgattc cgaagcccaa cctttcatag
     4861 aaggcggcgg tggaatcgaa atctcgtgat ggcaggttgg gcgtcgcttg gtcggtcatt
     4921 tcgaacccca gagtcccgct cagaagaact cgtcaagaag gcgatagaag gcgatgcgct
     4981 gcgaatcggg agcggcgata ccgtaaagca cgaggaagcg gtcagcccat tcgccgccaa
     5041 gctcttcagc aatatcacgg gtagccaacg ctatgtcctg atagcggtcc gccacaccca
     5101 gccggccaca gtcgatgaat ccagaaaagc ggccattttc caccatgata ttcggcaagc
     5161 aggcatcgcc atgggtcacg acgagatcct cgccgtcggg catgctcgcc ttgagcctgg
     5221 cgaacagttc ggctggcgcg agcccctgat gctcttcgtc cagatcatcc tgatcgacaa
     5281 gaccggcttc catccgagta cgtgctcgct cgatgcgatg tttcgcttgg tggtcgaatg
     5341 ggcaggtagc cggatcaagc gtatgcagcc gccgcattgc atcagccatg atggatactt
     5401 tctcggcagg agcaaggtga gatgacagga gatcctgccc cggcacttcg cccaatagca
     5461 gccagtccct tcccgcttca gtgacaacgt cgagcacagc tgcgcaagga acgcccgtcg
     5521 tggccagcca cgatagccgc gctgcctcgt cttgcagttc attcagggca ccggacaggt
     5581 cggtcttgac aaaaagaacc gggcgcccct gcgctgacag ccggaacacg gcggcatcag
     5641 agcagccgat tgtctgttgt gcccagtcat agccgaatag cctctccacc caagcggccg
     5701 gagaacctgc gtgcaatcca tcttgttcaa tcatgcgaaa cgatcctcat cctgtctctt
     5761 gatcgatctt tgcaaaagcc taggcctcca aaaaagcctc ctcactactt ctggaatagc
     5821 tcagaggccg aggcg
//
