LOCUS       sequence-75239-e        5536 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Express Venus and FLAG-tagged human CD59, a GPI-AP in mammalian 
            cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 5536)
  AUTHORS   Rivier AS, Castillon GA, Michon L, Fukasawa M, Romanova-Michaelides 
            M, Jaensch N, Hanada K, Watanabe R
  TITLE     Exit of GPI-anchored proteins from the ER differs in yeast and 
            mammalian cells.
  JOURNAL   Traffic. 2010 Aug;11(8):1017-33. doi: 
            10.1111/j.1600-0854.2010.01081.x. Epub 2010 May 11.
  PUBMED    20477992
REFERENCE   2  (bases 1 to 5536)
  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"; doi: 
            "10.1111/j.1600-0854.2010.01081.x"; journalName: "Traffic"; date: 
            "2010-08"; volume: "11"; issue: "8"; pages: "1017-33"
COMMENT     Sequence Label: pME-puro-Venus-FLAG-CD59
FEATURES             Location/Qualifiers
     source          1..5536
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     promoter        25..524
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(60..83)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     457..476
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             552..1151
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(552..571)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1048..1068
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(1683..1703)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        join(1688..2017,2018..2033,2034..2302)
                     /label=SR-alpha promoter
                     /note="hybrid promoter consisting of the SV40 early 
                     promoter plus part of the long terminal repeat of human 
                     T-lymphotrophic virus 1"
     rep_origin      1868..2003
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     LTR             2034..2302
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     intron          2332..2425
                     /label=SV40 intron
                     /note="modified SV40 intron with splice donor and acceptor 
                     sites"
     CDS             2555..2578
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     polyA_signal    2975..3109
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3012..3031)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3066..3085
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3148..3165)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3319..3907)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3399..3418)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4078..4869,4870..4938))
                     /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     4701..4720
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4939..5043)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5070..5525)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5207..5228)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5419..5438
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 aagcttcgac ctcgaaattc taccgggtag gggaggcgct tttcccaagg cagtctggag
       61 catgcgcttt agcagccccg ctgggcactt ggcgctacac aagtggcctc tggcctcgca
      121 cacattccac atccaccggt aggcgccaac cggctccgtt ctttggtggc cccttcgcgc
      181 caccttctac tcctccccta gtcaggaagt tcccccccgc cccgcagctc gcgtcgtgca
      241 ggacgtgaca aatggaagta gcacgtctca ctagtctcgt gcagatggac agcaccgctg
      301 agcaatggaa gcgggtaggc ctttggggca gcggccaata gcagctttgc tccttcgctt
      361 tctgggctca gaggctggga aggggtgggt ccgggggcgg gctcaggggc gggctcaggg
      421 gcggggcggg cgcccgaagg tcctccggag gcccggcatt ctgcacgctt caaaagcgca
      481 cgtctgccgc gctgttctcc tcttcctcat ctccgggcct ttcgacctgc atccatctag
      541 atcagcttac catgaccgag tacaagccca cggtgcgcct cgccacccgc gacgacgtcc
      601 ccagggccgt acgcaccctc gccgccgcgt tcgccgacta ccccgccacg cgccacaccg
      661 tcgatccgga ccgccacatc gagcgggtca ccgagctgca agaactcttc ctcacgcgcg
      721 tcgggctcga catcggcaag gtgtgggtcg cggacgacgg cgccgcggtg gcggtctgga
      781 ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga gatcggcccg cgcatggccg
      841 agttgagcgg ttcccggctg gccgcgcagc aacagatgga aggcctcctg gcgccgcacc
      901 ggcccaagga gcccgcgtgg ttcctggcca ccgtcggcgt ctcgcccgac caccagggca
      961 agggtctggg cagcgccgtc gtgctccccg gagtggaggc ggccgagcgc gccggggtgc
     1021 ccgccttcct ggagacctcc gcgccccgca acctcccctt ctacgagcgg ctcggcttca
     1081 ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac ctggtgcatg acccgcaagc
     1141 ccggtgcctg acgcccgccc cacgacccgc agcgcccgac cgaaaggagc gcacgacccc
     1201 atgcagggat gcagaaattg atgatctatt aaacaataaa gatgtccact aaaatggaag
     1261 tttttcctgt catactttgt taagaagggt gagaacagag tacctacatt ttgaatggaa
     1321 ggattggagc tacgggggtg ggggtggggt gggattagat aaatgcctgc tctttactga
     1381 aggctcttta ctattgcttt atgataatgt ttcatagttg gatatcataa tttaaacaag
     1441 caaaaccaaa ttaagggcca gctcattcct cccactcatg atctatagat ctatagatct
     1501 ctcgtgggat cattgttttt ctcttgattc ccactttgtg gttctaagta ctgtggtttc
     1561 caaatgtgtc agtttcatag cctgaagaac gagatcagca gcctctgttc cacatacact
     1621 tcattctcag tattgttttg ccaagttcta attccatcag aagctggtcg aagcttggct
     1681 gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     1741 gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc
     1801 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac
     1861 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     1921 aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta
     1981 gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctcctccgat cgaggggctc
     2041 gcatctctcc ttcacgcgcc cgccgcccta cctgaggccg ccatccacgc cggttgagtc
     2101 gcgttctgcc gcctcccgcc tgtggtgcct cctgaactgc gtccgccgtc taggtaagtt
     2161 taaagctcag gtcgagaccg ggcctttgtc cggcgctccc ttggagccta cctagactca
     2221 gccggctctc cacgctttgc ctgaccctgc ttgctcaact ctacgtcttt gtttcgtttt
     2281 ctgttctgcg ccgttacaga tccaagctct gaaaaaccag aaagttaact ggtaagttta
     2341 gtctttttgt cttttatttc aggtcccgga tccggtggtg caaatcaaag aactgctcct
     2401 cagtggatgt tgcctttact tctaggcctg tacggaagtg ttacttctgc tctaaaagct
     2461 gcggaattcc accatgggaa tccaaggagg gtctgtcctg ttcgggctgc tgctcgtcct
     2521 ggctgtcttc tgccattcag gtcatagcct gcaggactac aaggacgacg atgacaagct
     2581 tcagtgctac aactgtccta acccaactgc tgactgcaaa acagccgtca attgttcatc
     2641 tgattttgat gcgtgtctca ttaccaaagc tgggttacaa gtgtataaca agtgttggaa
     2701 gtttgagcat tgcaatttca acgacgtcac aacccgcttg agggaaaatg agctaacgta
     2761 ctactgctgc aagaaggacc tgtgtaactt taacgaacag ctcgagaatg gtgggacatc
     2821 cttatcagag aaaacagttc ttctgctggt gactccattt ctggcagcag cctggagcct
     2881 tcatccctaa gcggccgcta gactagtcta gagaaaaaac ctcccacacc tccccctgaa
     2941 cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg
     3001 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     3061 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctggatcc ccgggtaccg
     3121 agctcgaatt aattcctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     3181 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     3241 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     3301 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     3361 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     3421 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     3481 ctttctccct tcgggaagcg tggcgctttc tcaatgctca cgctgtaggt atctcagttc
     3541 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     3601 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     3661 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     3721 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     3781 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     3841 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     3901 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     3961 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     4021 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     4081 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     4141 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     4201 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     4261 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     4321 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     4381 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     4441 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     4501 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     4561 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     4621 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     4681 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     4741 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     4801 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     4861 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     4921 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     4981 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     5041 gcgcacattt ccccgaaaag tgccacctga aattgtaaac gttaatattt tgttaaaatt
     5101 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat
     5161 cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa
     5221 gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg
     5281 cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa
     5341 agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc
     5401 gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag
     5461 tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg
     5521 cgcgtcccat tcgcca
//
