LOCUS       sequence-417401-        8317 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 8317)
  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 8317)
  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-FCGRT-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8317
                     /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             2156..2191
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             2201..2224
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             3884..3928
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             3929..3955
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(4095..4112)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4101..4212
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4330..4350)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4335..4664
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4515..4650
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4577..4596
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4712..4759
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4778..5377
                     /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(4778..4797)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5274..5294
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5507..5628
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5544..5563)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5598..5617
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(5681..6472,6473..6541))
                     /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     6304..6323
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6542..6633)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6675..6694
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6700..6719
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      6805..7393
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7294..7313
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7547..7564
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     7662..7679
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7663..7679
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    8135..8151
                     /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 atctgctgaa tctcacctga gcctgctgta tcatctgacc gccgtgtcta gccccgctcc
     1381 tggaacacca gctttttggg tttccggctg gctgggccct cagcagtacc tgagctacaa
     1441 tagcctgaga ggcgaggccg aaccttgtgg cgcttgggtc tgggaaaatc aggtgtcctg
     1501 gtactgggag aaagagacaa ccgacctgcg gatcaaagag aagctgttcc tggaagcctt
     1561 caaggccctc ggcggaaagg gcccatatac actgcaaggc ctgctgggct gtgaactggg
     1621 ccctgataat accagcgtgc ccaccgccaa gttcgccctg aatggggaag agttcatgaa
     1681 tttcgacctg aagcaaggca cctggggcgg agattggcct gaagctctgg ccattagcca
     1741 gagatggcag cagcaggata aggccgccaa caaagagctg accttcctgc tgttcagctg
     1801 ccctcacaga ctgagagagc acctggaaag aggcagaggc aacctggaat ggaaagaacc
     1861 tcctagcatg cggctgaagg ccagacctag cagccctggc tttagcgtgc tgacctgtag
     1921 cgccttcagc ttctaccctc ctgaactcca gctgcggttc ctgagaaatg gactggccgc
     1981 tggaacaggc cagggcgatt ttggccctaa tagcgacggc agctttcacg cctctagcag
     2041 cctgacagtg aagtccggcg acgagcacca ctactgctgc attgtgcagc atgccggact
     2101 ggcccagcct ctgagagtgg aacttgagag ccctgccaag tcctctggcg cgccggaaga
     2161 tcaagttgat ccaagattga ttgatggcaa gggcggatct ctggaggtgc tgttccaggg
     2221 accaggcgga agcggcaatc ccggcggcga tctggccccc cagatgctgc gcgagctgca
     2281 ggagaccaat gccgccctgc aggacgtgcg cgagctgctg cgccaccgcg tgaaggagat
     2341 caccttcctg aagaacaccg tgatggagtg cgacgcctgc ggtatgcagc cctctagagg
     2401 ctccggctcc atcatcccag ttgaggagga gaacccggac ttctggaacc gcgaggcagc
     2461 cgaggccctg ggtgccgcca agaagctgca gcctgcacag acagccgcca agaacctcat
     2521 catcttcctg ggcgatggga tgggggtgtc tacggtgaca gctgccagga tcctaaaagg
     2581 gcagaagaag gacaaactgg ggcctgagat acccctggcc atggaccgct tcccatatgt
     2641 ggctctgtcc aagacataca atgtagacaa acatgtgcca gacagtggag ccacagccac
     2701 ggcctacctg tgcggggtca agggcaactt ccagaccatt ggcttgagtg cagccgcccg
     2761 ctttaaccag tgcaacacga cacgcggcaa cgaggtcatc tccgtgatga atcgggccaa
     2821 gaaagcaggg aagtcagtgg gagtggtaac caccacacga gtgcagcacg cctcgccagc
     2881 cggcacctac gcccacacgg tgaaccgcaa ctggtactcg gacgccgacg tgcctgcctc
     2941 ggcccgccag gaggggtgcc aggacatcgc tacgcagctc atctccaaca tggacattga
     3001 cgtgatccta ggtggaggcc gaaagtacat gtttcgcatg ggaaccccag accctgagta
     3061 cccagatgac tacagccaag gtgggaccag gctggacggg aagaatctgg tgcaggaatg
     3121 gctggcgaag cgccagggtg cccggtatgt gtggaaccgc actgagctca tgcaggcttc
     3181 cctggacccg tctgtgaccc atctcatggg tctctttgag cctggagaca tgaaatacga
     3241 gatccaccga gactccacac tggacccctc cctgatggag atgacagagg ctgccctgcg
     3301 cctgctgagc aggaaccccc gcggcttctt cctcttcgtg gagggtggtc gcatcgacca
     3361 tggtcatcat gaaagcaggg cttaccgggc actgactgag acgatcatgt tcgacgacgc
     3421 cattgagagg gcgggccagc tcaccagcga ggaggacacg ctgagcctcg tcactgccga
     3481 ccactcccac gtcttctcct tcggaggcta ccccctgcga gggagctcca tcttcgggct
     3541 ggcccctggc aaggcccggg acaggaaggc ctacacggtc ctcctatacg gaaacggtcc
     3601 aggctatgtg ctcaaggacg gcgcccggcc ggatgttacc gagagcgaga gcgggagccc
     3661 cgagtatcgg cagcagtcag cagtgcccct ggacgaagag acccacgcag gcgaggacgt
     3721 ggcggtgttc gcgcgcggcc cgcaggcgca cctggttcac ggcgtgcagg agcagacctt
     3781 catagcgcac gtcatggcct tcgccgcctg cctggagccc tacaccgcct gcgacctggc
     3841 gccccccgcc ggcaccaccg acgccgcgca cccgggtggc ggaggactga acgacatctt
     3901 cgaggctcag aaaatcgaat ggcacgaaca tcaccatcac catcaccatc accattaagg
     3961 ccggcctgta gtcactggca caacgcgtta cccggtaagc caatcgggta tacacggtcg
     4021 tcatactgca gacagggttc ttctactttg caagatagtc ttgagtagta aaataaatag
     4081 atagagaaaa tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
     4141 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     4201 ggaaattgca tcacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     4261 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     4321 taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     4381 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     4441 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     4501 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     4561 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc
     4621 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt
     4681 gtatatccat tttcggatct gatcagcacg tgttgacaat taatcatcgg catagtatat
     4741 cggcatagta taatacgaca aggtgaggaa ctaaaccatg accgagtaca agcccacggt
     4801 gcgcctcgcc acccgcgacg acgtccccag ggccgtacgc accctcgccg ccgcgttcgc
     4861 cgactacccc gccacgcgcc acaccgtcga tccggaccgc cacatcgagc gggtcaccga
     4921 gctgcaagaa ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga
     4981 cgacggcgcc gcggtggcgg tctggaccac gccggagagc gtcgaagcgg gggcggtgtt
     5041 cgccgagatc ggcccgcgca tggccgagtt gagcggttcc cggctggccg cgcagcaaca
     5101 gatggaaggc ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc tggccaccgt
     5161 cggcgtctcg cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc tccccggagt
     5221 ggaggcggcc gagcgcgccg gggtgcccgc cttcctggag acatccgcgc cccgcaacct
     5281 ccccttctac gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc
     5341 gcgcacctgg tgcatgaccc gcaagcccgg tgcctgacac gtgctacgag atttcgattc
     5401 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     5461 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc
     5521 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     5581 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat
     5641 accgtcgacc tctagctaga gcttggcgta atcatggtca ttaccaatgc ttaatcagtg
     5701 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     5761 tgtagataac tacgatacgg gagggcttac catctggccc cagcgctgcg atgataccgc
     5821 gagaaccacg ctcaccggct ccggatttat cagcaataaa ccagccagcc ggaagggccg
     5881 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     5941 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc atcgctacag
     6001 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     6061 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     6121 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     6181 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     6241 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     6301 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     6361 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     6421 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     6481 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     6541 tattcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     6601 acatatttga atgtatttag aaaaataaac aaataggggt cagtgttaca accaattaac
     6661 caattctgaa cattatcgcg agcccattta tacctgaata tggctcataa caccccttgc
     6721 tcatgaccaa aatcccttaa cgtgagttac gcgcgcgtcg ttccactgag cgtcagaccc
     6781 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     6841 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     6901 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     6961 gtagccgtag ttagcccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     7021 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     7081 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     7141 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     7201 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     7261 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     7321 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     7381 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     7441 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     7501 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     7561 cgaggaagcg gaaggcgaga gtagggaact gccaggcatc aaactaagca gaaggcccct
     7621 gacggatggc ctttttgcgt ttctacaaac tctttctgtg ttgtaaaacg acggccagtc
     7681 ttaagctcgg gccccctggg cggttctgat aacgagtaat cgttaatccg caaataacgt
     7741 aaaaacccgc ttcggcgggt ttttttatgg ggggagttta gggaaagagc atttgtcaga
     7801 atatttaagg gcgcctgtca ctttgcttga tatatgagaa ttatttaacc ttataaatga
     7861 gaaaaaagca acgcacttta aataagatac gttgcttttt cgattgatga acacctataa
     7921 ttaaactatt catctattat ttatgatttt ttgtatatac aatatttcta gtttgttaaa
     7981 gagaattaag aaaataaatc tcgaaaataa taaagggaaa atcagttttt gatatcaaaa
     8041 ttatacatgt caacgataat acaaaatata atacaaacta taagatgtta tcagtattta
     8101 ttatcattta gaataaattt tgtgtcgccc ttaattgtga gcggataaca attacgagct
     8161 tcatgcacag tggcgttgac tcaatattgg ccattagcca tattattcat tggttatata
     8221 gcataaatca atattggcta ttggccattg catacgttgt atctatatca taatatgtac
     8281 atttatattg gctcatgtcc aatatgaccg ccatgtt
//
