LOCUS       sequence-424407-        8347 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of cell-surface protein extracellular domain 
            fused to COMP5AP-AviTag-9xHis. Protein is secreted from cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8347)
  AUTHORS   Wojtowicz WM, Vielmetter J, Fernandes RA, Siepe DH, Eastman CL, 
            Chisholm GB, Cox S, Klock H, Anderson PW, Rue SM, Miller JJ, Glaser 
            SM, Bragstad ML, Vance J, Lam AW, Lesley SA, Zinn K, Garcia KC
  TITLE     A Human IgSF Cell-Surface Interactome Reveals a Complex Network of 
            Protein-Protein Interactions.
  JOURNAL   Cell. 2020 Aug 20;182(4):1027-1043.e17. doi: 
            10.1016/j.cell.2020.07.025.
  PUBMED    32822567
REFERENCE   2  (bases 1 to 8347)
  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: 
            "10.1016/j.cell.2020.07"; journalName: "Cell"; date: "2020-08-20- 
            20"; volume: "182"; issue: "4"; pages: "1027-1043.e17"
COMMENT     Sequence Label: pD649-HAsp-HLA-G-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8347
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        77..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     581..605
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        815..833
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     815..832
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     846..865
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             2186..2221
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             2231..2254
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             3914..3958
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             3959..3985
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(4125..4142)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4131..4242
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4360..4380)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4365..4694
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4545..4680
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4607..4626
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4742..4789
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4808..5407
                     /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(4808..4827)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5304..5324
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5537..5658
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5574..5593)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5628..5647
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(5711..6502,6503..6571))
                     /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     6334..6353
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6572..6663)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6705..6724
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6730..6749
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      6835..7423
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7324..7343
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7577..7594
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     7692..7709
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7693..7709
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    8165..8181
                     /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)."
ORIGIN
        1 ggcattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
      601 agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagacac cgggaccgat
      661 ccagcctccg cggccgggaa cggtgcattg gaacgcggat tccccgtgcc aagagtgacg
      721 taagtaccgc ctatagagtc tataggccca cccccttggc ttcgttagaa cgcggctaca
      781 attaatacat aaccttatgt atcatacaca tacgatttag gtgacactat agaataacat
      841 ccactttgcc tttctctcca caggtgtcca ctcccaggtc caactgcacc tcggttctat
      901 cgaaaacgcg cctgctagcg ccaccatgaa gaccatcatt gccctgagct acatcttctg
      961 cctggtgttc gctgcggccg ccattcagag aacccctaag atccaggtgt acagcagaca
     1021 ccccgccgag aacggcaaga gcaacttcct gaactgctac gtgtccggct ttcaccccag
     1081 cgacattgag gtggacctgc tgaagaacgg cgagcggatc gagaaggtgg aacacagcga
     1141 tctgagcttc agcaaggact ggtccttcta cctgctgtac tacaccgagt tcacccctac
     1201 cgagaaggac gagtacgcct gcagagtgaa ccacgtgaca ctgagccagc ctaagatcgt
     1261 gaagtgggac agagacatgg gcggaggcgg atctggtggc ggaggaagtg gcggcggagg
     1321 atctggatct cacagcatga gatacttctc tgccgccgtg tccagacctg gaagaggcga
     1381 gcctagattc attgccatgg gctacgtgga cgacacccag ttcgtcagat tcgacagcga
     1441 cagcgcctgt cctcggatgg aacctagagc accttgggtc gagcaagagg gccctgagta
     1501 ctgggaagaa gagacacgga acaccaaggc tcacgcccag accgacagaa tgaacctgca
     1561 gaccctgcgg ggctactaca atcagtctga ggccagcagc catactctgc agtggatgat
     1621 cggctgcgat ctgggctctg atggcagact gctgagaggc tacgagcagt acgcctacga
     1681 cggcaaggat tatctggccc tgaacgagga cctgcggtct tggacagctg ccgatacagc
     1741 cgctcagatc agcaagagaa agtgcgaggc cgccaatgtg gccgaacaga gaagggctta
     1801 cctggaaggc acctgtgtgg aatggctgca cagatacctg gaaaacggca aagagatgct
     1861 gcagcgggcc gatcctccta agacacatgt gacccaccat cctgtgttcg actacgaggc
     1921 cacactgaga tgttgggccc tgggctttta ccctgccgag atcatcctga cctggcagag
     1981 agatggcgag gatcagaccc aggacgtgga actggtggaa acaagacctg ccggcgacgg
     2041 caccttccag aaatgggctg ctgtggtggt tcccagcggc gaggaacaga gatacacctg
     2101 tcacgtgcag cacgagggac tgcctgaacc tctgatgctg agatggaagc agagcagcct
     2161 gcctacaatc cccattggcg cgccggaaga tcaagttgat ccaagattga ttgatggcaa
     2221 gggcggatct ctggaggtgc tgttccaggg accaggcgga agcggcaatc ccggcggcga
     2281 tctggccccc cagatgctgc gcgagctgca ggagaccaat gccgccctgc aggacgtgcg
     2341 cgagctgctg cgccaccgcg tgaaggagat caccttcctg aagaacaccg tgatggagtg
     2401 cgacgcctgc ggtatgcagc cctctagagg ctccggctcc atcatcccag ttgaggagga
     2461 gaacccggac ttctggaacc gcgaggcagc cgaggccctg ggtgccgcca agaagctgca
     2521 gcctgcacag acagccgcca agaacctcat catcttcctg ggcgatggga tgggggtgtc
     2581 tacggtgaca gctgccagga tcctaaaagg gcagaagaag gacaaactgg ggcctgagat
     2641 acccctggcc atggaccgct tcccatatgt ggctctgtcc aagacataca atgtagacaa
     2701 acatgtgcca gacagtggag ccacagccac ggcctacctg tgcggggtca agggcaactt
     2761 ccagaccatt ggcttgagtg cagccgcccg ctttaaccag tgcaacacga cacgcggcaa
     2821 cgaggtcatc tccgtgatga atcgggccaa gaaagcaggg aagtcagtgg gagtggtaac
     2881 caccacacga gtgcagcacg cctcgccagc cggcacctac gcccacacgg tgaaccgcaa
     2941 ctggtactcg gacgccgacg tgcctgcctc ggcccgccag gaggggtgcc aggacatcgc
     3001 tacgcagctc atctccaaca tggacattga cgtgatccta ggtggaggcc gaaagtacat
     3061 gtttcgcatg ggaaccccag accctgagta cccagatgac tacagccaag gtgggaccag
     3121 gctggacggg aagaatctgg tgcaggaatg gctggcgaag cgccagggtg cccggtatgt
     3181 gtggaaccgc actgagctca tgcaggcttc cctggacccg tctgtgaccc atctcatggg
     3241 tctctttgag cctggagaca tgaaatacga gatccaccga gactccacac tggacccctc
     3301 cctgatggag atgacagagg ctgccctgcg cctgctgagc aggaaccccc gcggcttctt
     3361 cctcttcgtg gagggtggtc gcatcgacca tggtcatcat gaaagcaggg cttaccgggc
     3421 actgactgag acgatcatgt tcgacgacgc cattgagagg gcgggccagc tcaccagcga
     3481 ggaggacacg ctgagcctcg tcactgccga ccactcccac gtcttctcct tcggaggcta
     3541 ccccctgcga gggagctcca tcttcgggct ggcccctggc aaggcccggg acaggaaggc
     3601 ctacacggtc ctcctatacg gaaacggtcc aggctatgtg ctcaaggacg gcgcccggcc
     3661 ggatgttacc gagagcgaga gcgggagccc cgagtatcgg cagcagtcag cagtgcccct
     3721 ggacgaagag acccacgcag gcgaggacgt ggcggtgttc gcgcgcggcc cgcaggcgca
     3781 cctggttcac ggcgtgcagg agcagacctt catagcgcac gtcatggcct tcgccgcctg
     3841 cctggagccc tacaccgcct gcgacctggc gccccccgcc ggcaccaccg acgccgcgca
     3901 cccgggtggc ggaggactga acgacatctt cgaggctcag aaaatcgaat ggcacgaaca
     3961 tcaccatcac catcaccatc accattaagg ccggcctgta gtcactggca caacgcgtta
     4021 cccggtaagc caatcgggta tacacggtcg tcatactgca gacagggttc ttctactttg
     4081 caagatagtc ttgagtagta aaataaatag atagagaaaa tcagcctcga ctgtgccttc
     4141 tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc
     4201 cactcccact gtcctttcct aataaaatga ggaaattgca tcacaacact caaccctatc
     4261 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     4321 gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt
     4381 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     4441 cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     4501 atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc
     4561 cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg
     4621 ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
     4681 taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct gatcagcacg
     4741 tgttgacaat taatcatcgg catagtatat cggcatagta taatacgaca aggtgaggaa
     4801 ctaaaccatg accgagtaca agcccacggt gcgcctcgcc acccgcgacg acgtccccag
     4861 ggccgtacgc accctcgccg ccgcgttcgc cgactacccc gccacgcgcc acaccgtcga
     4921 tccggaccgc cacatcgagc gggtcaccga gctgcaagaa ctcttcctca cgcgcgtcgg
     4981 gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg tctggaccac
     5041 gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca tggccgagtt
     5101 gagcggttcc cggctggccg cgcagcaaca gatggaaggc ctcctggcgc cgcaccggcc
     5161 caaggagccc gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc agggcaaggg
     5221 tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg gggtgcccgc
     5281 cttcctggag acatccgcgc cccgcaacct ccccttctac gagcggctcg gcttcaccgt
     5341 caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc gcaagcccgg
     5401 tgcctgacac gtgctacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
     5461 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     5521 gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     5581 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     5641 actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta
     5701 atcatggtca ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     5761 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     5821 catctggccc cagcgctgcg atgataccgc gagaaccacg ctcaccggct ccggatttat
     5881 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     5941 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     6001 gtttgcgcaa cgttgttgcc atcgctacag gcatcgtggt gtcacgctcg tcgtttggta
     6061 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     6121 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     6181 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     6241 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     6301 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     6361 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     6421 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     6481 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     6541 taagggcgac acggaaatgt tgaatactca tattcttcct ttttcaatat tattgaagca
     6601 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     6661 aaataggggt cagtgttaca accaattaac caattctgaa cattatcgcg agcccattta
     6721 tacctgaata tggctcataa caccccttgc tcatgaccaa aatcccttaa cgtgagttac
     6781 gcgcgcgtcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     6841 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     6901 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     6961 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttagcccacc acttcaagaa
     7021 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     7081 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     7141 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     7201 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     7261 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     7321 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     7381 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     7441 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     7501 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     7561 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaaggcgaga gtagggaact
     7621 gccaggcatc aaactaagca gaaggcccct gacggatggc ctttttgcgt ttctacaaac
     7681 tctttctgtg ttgtaaaacg acggccagtc ttaagctcgg gccccctggg cggttctgat
     7741 aacgagtaat cgttaatccg caaataacgt aaaaacccgc ttcggcgggt ttttttatgg
     7801 ggggagttta gggaaagagc atttgtcaga atatttaagg gcgcctgtca ctttgcttga
     7861 tatatgagaa ttatttaacc ttataaatga gaaaaaagca acgcacttta aataagatac
     7921 gttgcttttt cgattgatga acacctataa ttaaactatt catctattat ttatgatttt
     7981 ttgtatatac aatatttcta gtttgttaaa gagaattaag aaaataaatc tcgaaaataa
     8041 taaagggaaa atcagttttt gatatcaaaa ttatacatgt caacgataat acaaaatata
     8101 atacaaacta taagatgtta tcagtattta ttatcattta gaataaattt tgtgtcgccc
     8161 ttaattgtga gcggataaca attacgagct tcatgcacag tggcgttgac tcaatattgg
     8221 ccattagcca tattattcat tggttatata gcataaatca atattggcta ttggccattg
     8281 catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc aatatgaccg
     8341 ccatgtt
//
