LOCUS       sequence-492992-        7636 bp    DNA     circular SYN 18-NOV-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 7636)
  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 7636)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 18, 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-EDAR-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..7636
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      24..159
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     86..105
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        221..268
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             287..886
                     /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(287..306)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     783..803
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    1016..1137
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1053..1072)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1107..1126
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(1190..1981,1982..2050))
                     /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     1813..1832
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2051..2142)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2184..2203
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2209..2228
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2314..2902
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2803..2822
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3056..3073
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     3171..3188
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3172..3188
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3644..3660
                     /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)."
     enhancer        3827..4206
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4207..4410
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4361..4381
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4407..4431
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        4641..4659
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     4641..4658
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     4672..4691
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             5346..5369
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             7029..7073
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             7074..7100
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(7240..7257)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    7246..7357
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(7475..7495)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        join(7480..7636,1..173)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
ORIGIN
        1 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
       61 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg
      121 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc
      181 cgggagcttg tatatccatt ttcggatctg atcagcacgt gttgacaatt aatcatcggc
      241 atagtatatc ggcatagtat aatacgacaa ggtgaggaac taaaccatga ccgagtacaa
      301 gcccacggtg cgcctcgcca cccgcgacga cgtccccagg gccgtacgca ccctcgccgc
      361 cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat ccggaccgcc acatcgagcg
      421 ggtcaccgag ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg gcaaggtgtg
      481 ggtcgcggac gacggcgccg cggtggcggt ctggaccacg ccggagagcg tcgaagcggg
      541 ggcggtgttc gccgagatcg gcccgcgcat ggccgagttg agcggttccc ggctggccgc
      601 gcagcaacag atggaaggcc tcctggcgcc gcaccggccc aaggagcccg cgtggttcct
      661 ggccaccgtc ggcgtctcgc ccgaccacca gggcaagggt ctgggcagcg ccgtcgtgct
      721 ccccggagtg gaggcggccg agcgcgccgg ggtgcccgcc ttcctggaga catccgcgcc
      781 ccgcaacctc cccttctacg agcggctcgg cttcaccgtc accgccgacg tcgaggtgcc
      841 cgaaggaccg cgcacctggt gcatgacccg caagcccggt gcctgacacg tgctacgaga
      901 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc
      961 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt
     1021 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     1081 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
     1141 tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat taccaatgct
     1201 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     1261 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agcgctgcga
     1321 tgataccgcg agaaccacgc tcaccggctc cggatttatc agcaataaac cagccagccg
     1381 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     1441 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     1501 tcgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     1561 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     1621 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     1681 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     1741 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     1801 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     1861 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     1921 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     1981 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     2041 gaatactcat attcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     2101 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtc agtgttacaa
     2161 ccaattaacc aattctgaac attatcgcga gcccatttat acctgaatat ggctcataac
     2221 accccttgct catgaccaaa atcccttaac gtgagttacg cgcgcgtcgt tccactgagc
     2281 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     2341 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     2401 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     2461 tcttctagtg tagccgtagt tagcccacca cttcaagaac tctgtagcac cgcctacata
     2521 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     2581 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     2641 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     2701 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     2761 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     2821 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     2881 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     2941 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     3001 tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga
     3061 gtcagtgagc gaggaagcgg aaggcgagag tagggaactg ccaggcatca aactaagcag
     3121 aaggcccctg acggatggcc tttttgcgtt tctacaaact ctttctgtgt tgtaaaacga
     3181 cggccagtct taagctcggg ccccctgggc ggttctgata acgagtaatc gttaatccgc
     3241 aaataacgta aaaacccgct tcggcgggtt tttttatggg gggagtttag ggaaagagca
     3301 tttgtcagaa tatttaaggg cgcctgtcac tttgcttgat atatgagaat tatttaacct
     3361 tataaatgag aaaaaagcaa cgcactttaa ataagatacg ttgctttttc gattgatgaa
     3421 cacctataat taaactattc atctattatt tatgattttt tgtatataca atatttctag
     3481 tttgttaaag agaattaaga aaataaatct cgaaaataat aaagggaaaa tcagtttttg
     3541 atatcaaaat tatacatgtc aacgataata caaaatataa tacaaactat aagatgttat
     3601 cagtatttat tatcatttag aataaatttt gtgtcgccct taattgtgag cggataacaa
     3661 ttacgagctt catgcacagt ggcgttgact caatattggc cattagccat attattcatt
     3721 ggttatatag cataaatcaa tattggctat tggccattgc atacgttgta tctatatcat
     3781 aatatgtaca tttatattgg ctcatgtcca atatgaccgc catgttggca ttgattattg
     3841 actagttatt aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc
     3901 cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca
     3961 ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt
     4021 caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg
     4081 ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag
     4141 tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt
     4201 accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg
     4261 ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa
     4321 cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt
     4381 gtacggtggg aggtctatat aagcagagct cgtttagtga accgtcagat cgcctggaga
     4441 cgccatccac gctgttttga cctccataga agacaccggg accgatccag cctccgcggc
     4501 cgggaacggt gcattggaac gcggattccc cgtgccaaga gtgacgtaag taccgcctat
     4561 agagtctata ggcccacccc cttggcttcg ttagaacgcg gctacaatta atacataacc
     4621 ttatgtatca tacacatacg atttaggtga cactatagaa taacatccac tttgcctttc
     4681 tctccacagg tgtccactcc caggtccaac tgcacctcgg ttctatcgaa aacgcgcctg
     4741 ctagcgccac catgaagacc atcattgccc tgagctacat cttctgcctg gtgttcgctg
     4801 cggccgccga gtatagcaat tgcggcgaga acgagtacta caaccagacc accggcctgt
     4861 gccaagagtg tcctccttgt ggacctggcg aggaacctta cctgagctgt ggctacggca
     4921 ccaaggatga ggattacggc tgtgtgcctt gtcctgccga gaagttcagc aaaggcggct
     4981 accagatctg cagacggcac aaggactgcg agggcttctt tagagccaca gtgctgaccc
     5041 ctggcgacat ggaaaatgat gccgagtgtg gaccttgcct gcctggctac tacatgctgg
     5101 aaaacagacc ccggaacatc tacggcatgg tctgctactc ttgtctgctg gcccctccta
     5161 acaccaaaga atgtgtggga gccacaagcg gcgccagcgc caattttcct ggcacaagcg
     5221 gaagcagcac actgagccct tttcagcacg cccacaaaga gctgtctgga cagggacatc
     5281 tggccaccgc tggcgcgccg gaagatcaag ttgatccaag attgattgat ggcaagggcg
     5341 gatctctgga ggtgctgttc cagggaccag gcggaagcgg caatcccggc ggcgatctgg
     5401 ccccccagat gctgcgcgag ctgcaggaga ccaatgccgc cctgcaggac gtgcgcgagc
     5461 tgctgcgcca ccgcgtgaag gagatcacct tcctgaagaa caccgtgatg gagtgcgacg
     5521 cctgcggtat gcagccctct agaggctccg gctccatcat cccagttgag gaggagaacc
     5581 cggacttctg gaaccgcgag gcagccgagg ccctgggtgc cgccaagaag ctgcagcctg
     5641 cacagacagc cgccaagaac ctcatcatct tcctgggcga tgggatgggg gtgtctacgg
     5701 tgacagctgc caggatccta aaagggcaga agaaggacaa actggggcct gagatacccc
     5761 tggccatgga ccgcttccca tatgtggctc tgtccaagac atacaatgta gacaaacatg
     5821 tgccagacag tggagccaca gccacggcct acctgtgcgg ggtcaagggc aacttccaga
     5881 ccattggctt gagtgcagcc gcccgcttta accagtgcaa cacgacacgc ggcaacgagg
     5941 tcatctccgt gatgaatcgg gccaagaaag cagggaagtc agtgggagtg gtaaccacca
     6001 cacgagtgca gcacgcctcg ccagccggca cctacgccca cacggtgaac cgcaactggt
     6061 actcggacgc cgacgtgcct gcctcggccc gccaggaggg gtgccaggac atcgctacgc
     6121 agctcatctc caacatggac attgacgtga tcctaggtgg aggccgaaag tacatgtttc
     6181 gcatgggaac cccagaccct gagtacccag atgactacag ccaaggtggg accaggctgg
     6241 acgggaagaa tctggtgcag gaatggctgg cgaagcgcca gggtgcccgg tatgtgtgga
     6301 accgcactga gctcatgcag gcttccctgg acccgtctgt gacccatctc atgggtctct
     6361 ttgagcctgg agacatgaaa tacgagatcc accgagactc cacactggac ccctccctga
     6421 tggagatgac agaggctgcc ctgcgcctgc tgagcaggaa cccccgcggc ttcttcctct
     6481 tcgtggaggg tggtcgcatc gaccatggtc atcatgaaag cagggcttac cgggcactga
     6541 ctgagacgat catgttcgac gacgccattg agagggcggg ccagctcacc agcgaggagg
     6601 acacgctgag cctcgtcact gccgaccact cccacgtctt ctccttcgga ggctaccccc
     6661 tgcgagggag ctccatcttc gggctggccc ctggcaaggc ccgggacagg aaggcctaca
     6721 cggtcctcct atacggaaac ggtccaggct atgtgctcaa ggacggcgcc cggccggatg
     6781 ttaccgagag cgagagcggg agccccgagt atcggcagca gtcagcagtg cccctggacg
     6841 aagagaccca cgcaggcgag gacgtggcgg tgttcgcgcg cggcccgcag gcgcacctgg
     6901 ttcacggcgt gcaggagcag accttcatag cgcacgtcat ggccttcgcc gcctgcctgg
     6961 agccctacac cgcctgcgac ctggcgcccc ccgccggcac caccgacgcc gcgcacccgg
     7021 gtggcggagg actgaacgac atcttcgagg ctcagaaaat cgaatggcac gaacatcacc
     7081 atcaccatca ccatcaccat taaggccggc ctgtagtcac tggcacaacg cgttacccgg
     7141 taagccaatc gggtatacac ggtcgtcata ctgcagacag ggttcttcta ctttgcaaga
     7201 tagtcttgag tagtaaaata aatagataga gaaaatcagc ctcgactgtg ccttctagtt
     7261 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     7321 ccactgtcct ttcctaataa aatgaggaaa ttgcatcaca acactcaacc ctatctcggt
     7381 ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct
     7441 gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga
     7501 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     7561 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     7621 aattagtcag caacca
//
