LOCUS       sequence-177916-        5477 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A mammalian expression plasmid expressing CD8a luminal and 
            transmembrane domains followed by cNLS (classical nuclear 
            localization signal) and C-terminus EGFP..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5477)
  AUTHORS   Madugula V, Lu L
  TITLE     A ternary complex comprising transportin1, Rab8 and the ciliary 
            targeting signal directs proteins to ciliary membranes.
  JOURNAL   J Cell Sci. 2016 Oct 15;129(20):3922-3934. Epub 2016 Sep 15.
  PUBMED    27633000
REFERENCE   2  (bases 1 to 5477)
  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: "2016-10-15"; volume: "129(20)"; pages: "3922-3934. 
            Epub 2016 Sep 15"
COMMENT     Sequence Label: CD8a-cNLS-EGFP
FEATURES             Location/Qualifiers
     source          1..5477
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      10..598
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     499..518
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        773..1076
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1077..1280
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1231..1251
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             2039..2059
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2063..2083
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     regulatory      2129..2138
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2135..2137,2138..2140,2141..2854)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2135..2137,2138..2140,2141..2851)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2180..2201)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2441..2460)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2788..2809
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2977..3098
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3014..3033)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3068..3087
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3105..3560)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3242..3263)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3454..3473
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3587..3691
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3693..4050
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3693..3884
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3716..3736)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3901..4036
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4085..4879
                     /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             4085..4879
                     /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(4139..4158)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4749..4768
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5067..5086)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5111..5158
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
ORIGIN
        1 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
       61 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
      121 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg
      181 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
      241 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
      301 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
      361 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
      421 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
      481 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
      541 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
      601 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
      661 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgccatgc attagttatt
      721 aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat
      781 aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa
      841 taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg
      901 agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc
      961 cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct
     1021 tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga
     1081 tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa
     1141 gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc
     1201 caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg
     1261 aggtctatat aagcagagct ggtttagtga accgtcagat ccgctagcgc taccggactc
     1321 agatctcgag atggccttac cagtgaccgc cttgctcctg ccgctggcct tgctgctcca
     1381 cgccgccagg ccgagccagt tccgggtgtc gccgctggat cggacctgga acctgggcga
     1441 gacagtggag ctgaagtgcc aggtgctgct gtccaacccg acgtcgggct gctcgtggct
     1501 cttccagccg cgcggcgccg ccgccagtcc caccttcctc ctatacctct cccaaaacaa
     1561 gcccaaggcg gccgaggggc tggacaccca gcggttctcg ggcaagaggt tgggggacac
     1621 cttcgtcctc accctgagcg acttccgccg agagaacgag ggctactatt tctgctcggc
     1681 cctgagcaac tccatcatgt acttcagcca cttcgtgccg gtcttcctgc cagcgaagcc
     1741 caccacgacg ccagcgccgc gaccaccaac accggcgccc accatcgcgt cgcagcccct
     1801 gtccctgcgc ccagaggcgt gccggccagc ggcggggggc gcagtgcaca cgagggggct
     1861 ggacttcgcc tgtgatatct acatctgggc gcccttggcc gggacttgtg gggtccttct
     1921 cctgtcactg gttatcaccc tttactgcaa ccacaggaac cgaagacgtg tttgcaaatg
     1981 tccccggcct gtggtcaaat cgggagacaa gcccagcctt tcggcgagat acgtcgatcc
     2041 aaaaaagaag agaaaggtag atccaaaaaa gaagagaaag gtaggaattc tgcagtcgac
     2101 ggtaccgcgg gcccgggatc caccggtcgc caccatggtg agcaagggcg aggagctgtt
     2161 caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag
     2221 cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg
     2281 caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctga cctacggcgt
     2341 gcagtgcttc agccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat
     2401 gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac
     2461 ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat
     2521 cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca
     2581 caacgtctat atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg
     2641 ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat
     2701 cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcacccagt ccgccctgag
     2761 caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg
     2821 gatcactctc ggcatggacg agctgtacaa gtaaagcggc cgcgactcta gatcataatc
     2881 agccatacca catttgtaga ggttttactt gctttaaaaa acctcccaca cctccccctg
     2941 aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat
     3001 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     3061 tctagttgtg gtttgtccaa actcatcaat gtatcttaag gcgtaaattg taagcgttaa
     3121 tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta accaataggc
     3181 cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt tgagtgttgt
     3241 tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca aagggcgaaa
     3301 aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa gttttttggg
     3361 gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat ttagagcttg
     3421 acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc
     3481 tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa
     3541 tgcgccgcta cagggcgcgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     3601 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     3661 aatgcttcaa taatattgaa aaaggaagag tcctgaggcg gaaagaacca gctgtggaat
     3721 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     3781 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     3841 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     3901 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     3961 tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga
     4021 ggcttttttg gaggcctagg cttttgcaaa gatcgatcaa gagacaggat gaggatcgtt
     4081 tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct
     4141 attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct
     4201 gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga
     4261 actgcaagac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc
     4321 tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg
     4381 gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc
     4441 aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca
     4501 tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga
     4561 cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg cgagcatgcc
     4621 cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga
     4681 aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca
     4741 ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg
     4801 cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct
     4861 tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc
     4921 aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga
     4981 atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc
     5041 ttcgcccacc ctagggggag gctaactgaa acacggaagg agacaatacc ggaaggaacc
     5101 cgcgctatga cggcaataaa aagacagaat aaaacgcacg gtgttgggtc gtttgttcat
     5161 aaacgcgggg ttcggtccca gggctggcac tctgtcgata ccccaccgag accccattgg
     5221 ggccaatacg cccgcgtttc ttccttttcc ccaccccacc ccccaagttc gggtgaaggc
     5281 ccagggctcg cagccaacgt cggggcggca ggccctgcca tagcctcagg ttactcatat
     5341 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     5401 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     5461 cccgtagaaa agatcaa
//
