LOCUS       sequence-385995-        8887 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 8887)
  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 8887)
  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-SEMA6B-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8887
                     /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             2726..2761
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             2771..2794
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             4454..4498
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             4499..4525
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(4665..4682)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4671..4782
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4900..4920)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4905..5234
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5085..5220
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5147..5166
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        5282..5329
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5348..5947
                     /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(5348..5367)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5844..5864
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6077..6198
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6114..6133)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6168..6187
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(6251..7042,7043..7111))
                     /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     6874..6893
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7112..7203)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7245..7264
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     7270..7289
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      7375..7963
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7864..7883
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8117..8134
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     8232..8249
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     8233..8249
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    8705..8721
                     /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 ccttgtttcc tgaagaacct cctcctctgt ctgtggcccc
     1021 tcgggactac ctgaatcact accctgtgtt tgttggcagc ggccctggta gacttacacc
     1081 agctgaaggc gccgacgacc tgaatatcca gcgggtgctg agagtgaaca gaaccctgtt
     1141 catcggcgac cgggacaacc tgtacagagt ggaactggaa cctcctacca gcaccgagct
     1201 gagataccag agaaagctga cctggcggag caaccccagc gacatcaacg tgtgcagaat
     1261 gaagggcaag caagagggcg agtgccggaa ctttgtgaag gtgctgctgc tgagagatga
     1321 gagcacactg ttcgtgtgcg gcagcaacgc cttcaatccc gtgtgcgcca actacagcat
     1381 cgacacactg cagcccgtgg gcgacaatat ctctggcatg gccagatgtc cctacgatcc
     1441 caaacatgcc aacgtggccc tgttcagcga cggcatgctg tttaccgcca ccgtgaccga
     1501 ctttctggcc atcgatgccg tgatctacag aagcctgggc gacagaccta cactgagaac
     1561 cgtgaagcac gacagcaagt ggttcaaaga accctacttc gtgcacgccg tggaatgggg
     1621 cagccacgtg tactttttct tccgggaaat cgccatggaa ttcaactacc tggaaaaggt
     1681 ggtggtgtcc agagtggcca gagtgtgcaa aaacgatgtc ggcggcagcc ccagagtgct
     1741 ggaaaagcag tggacaagct tcctgaaggc ccggctgaat tgctctgtgc ctggcgacag
     1801 ccacttctac ttcaatgtgc tgcaggccgt gaccggcgtg gtgtctcttg gaggtagacc
     1861 tgtggtgctg gccgtgttta gcacccctag caattctatc cctggcagcg ccgtgtgtgc
     1921 cttcgatctg acacaagtgg ccgccgtgtt cgagggcaga ttcagagagc agaagtcccc
     1981 tgagagcatc tggacccctg tgcctgaaga tcaggtgcca agacctagac ctggctgttg
     2041 tgctgcccct ggcatgcagt acaatgcctc ttctgccctg cctgacgaca tcctgaactt
     2101 cgtgaaaaca caccctctga tggacgaggc cgtgccttct cttggacatg ccccttggat
     2161 tctgcggacc ctgatgagac atcagctgac aagagtggcc gtggatgttg gagccggacc
     2221 ttggggaaat cagaccgtgg tgtttctggg ctctgaggcc ggcacagtgc tgaaatttct
     2281 tgtgcggcct aatgccagca caagcggaac aagcggcctg agcgtgttcc tggaagagtt
     2341 cgagacatac agacccgaca gatgcggtag accaggcggc ggagaaacag gacagagact
     2401 gctgtctctt gagctggatg ctgcttctgg tggcctgctg gccgcctttc ctagatgtgt
     2461 tgttagagtg cctgtggcca ggtgccagca gtacagcggc tgcatgaaga actgcatcgg
     2521 ctctcaggac ccttactgtg gatgggcccc tgacggcagc tgtatctttc tgtctcctgg
     2581 caccagagcc gccttcgaac aagatgtgtc tggcgcctct acaagcggac tgggagattg
     2641 cacaggactg ctgagggcca gcctgtctga agatagagct ggcctggtgt ccgtgaacct
     2701 gctggtcaca tcttctggcg cgccggaaga tcaagttgat ccaagattga ttgatggcaa
     2761 gggcggatct ctggaggtgc tgttccaggg accaggcgga agcggcaatc ccggcggcga
     2821 tctggccccc cagatgctgc gcgagctgca ggagaccaat gccgccctgc aggacgtgcg
     2881 cgagctgctg cgccaccgcg tgaaggagat caccttcctg aagaacaccg tgatggagtg
     2941 cgacgcctgc ggtatgcagc cctctagagg ctccggctcc atcatcccag ttgaggagga
     3001 gaacccggac ttctggaacc gcgaggcagc cgaggccctg ggtgccgcca agaagctgca
     3061 gcctgcacag acagccgcca agaacctcat catcttcctg ggcgatggga tgggggtgtc
     3121 tacggtgaca gctgccagga tcctaaaagg gcagaagaag gacaaactgg ggcctgagat
     3181 acccctggcc atggaccgct tcccatatgt ggctctgtcc aagacataca atgtagacaa
     3241 acatgtgcca gacagtggag ccacagccac ggcctacctg tgcggggtca agggcaactt
     3301 ccagaccatt ggcttgagtg cagccgcccg ctttaaccag tgcaacacga cacgcggcaa
     3361 cgaggtcatc tccgtgatga atcgggccaa gaaagcaggg aagtcagtgg gagtggtaac
     3421 caccacacga gtgcagcacg cctcgccagc cggcacctac gcccacacgg tgaaccgcaa
     3481 ctggtactcg gacgccgacg tgcctgcctc ggcccgccag gaggggtgcc aggacatcgc
     3541 tacgcagctc atctccaaca tggacattga cgtgatccta ggtggaggcc gaaagtacat
     3601 gtttcgcatg ggaaccccag accctgagta cccagatgac tacagccaag gtgggaccag
     3661 gctggacggg aagaatctgg tgcaggaatg gctggcgaag cgccagggtg cccggtatgt
     3721 gtggaaccgc actgagctca tgcaggcttc cctggacccg tctgtgaccc atctcatggg
     3781 tctctttgag cctggagaca tgaaatacga gatccaccga gactccacac tggacccctc
     3841 cctgatggag atgacagagg ctgccctgcg cctgctgagc aggaaccccc gcggcttctt
     3901 cctcttcgtg gagggtggtc gcatcgacca tggtcatcat gaaagcaggg cttaccgggc
     3961 actgactgag acgatcatgt tcgacgacgc cattgagagg gcgggccagc tcaccagcga
     4021 ggaggacacg ctgagcctcg tcactgccga ccactcccac gtcttctcct tcggaggcta
     4081 ccccctgcga gggagctcca tcttcgggct ggcccctggc aaggcccggg acaggaaggc
     4141 ctacacggtc ctcctatacg gaaacggtcc aggctatgtg ctcaaggacg gcgcccggcc
     4201 ggatgttacc gagagcgaga gcgggagccc cgagtatcgg cagcagtcag cagtgcccct
     4261 ggacgaagag acccacgcag gcgaggacgt ggcggtgttc gcgcgcggcc cgcaggcgca
     4321 cctggttcac ggcgtgcagg agcagacctt catagcgcac gtcatggcct tcgccgcctg
     4381 cctggagccc tacaccgcct gcgacctggc gccccccgcc ggcaccaccg acgccgcgca
     4441 cccgggtggc ggaggactga acgacatctt cgaggctcag aaaatcgaat ggcacgaaca
     4501 tcaccatcac catcaccatc accattaagg ccggcctgta gtcactggca caacgcgtta
     4561 cccggtaagc caatcgggta tacacggtcg tcatactgca gacagggttc ttctactttg
     4621 caagatagtc ttgagtagta aaataaatag atagagaaaa tcagcctcga ctgtgccttc
     4681 tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc
     4741 cactcccact gtcctttcct aataaaatga ggaaattgca tcacaacact caaccctatc
     4801 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     4861 gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt
     4921 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     4981 cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     5041 atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc
     5101 cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg
     5161 ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
     5221 taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct gatcagcacg
     5281 tgttgacaat taatcatcgg catagtatat cggcatagta taatacgaca aggtgaggaa
     5341 ctaaaccatg accgagtaca agcccacggt gcgcctcgcc acccgcgacg acgtccccag
     5401 ggccgtacgc accctcgccg ccgcgttcgc cgactacccc gccacgcgcc acaccgtcga
     5461 tccggaccgc cacatcgagc gggtcaccga gctgcaagaa ctcttcctca cgcgcgtcgg
     5521 gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg tctggaccac
     5581 gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca tggccgagtt
     5641 gagcggttcc cggctggccg cgcagcaaca gatggaaggc ctcctggcgc cgcaccggcc
     5701 caaggagccc gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc agggcaaggg
     5761 tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg gggtgcccgc
     5821 cttcctggag acatccgcgc cccgcaacct ccccttctac gagcggctcg gcttcaccgt
     5881 caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc gcaagcccgg
     5941 tgcctgacac gtgctacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
     6001 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     6061 gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     6121 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     6181 actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta
     6241 atcatggtca ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     6301 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     6361 catctggccc cagcgctgcg atgataccgc gagaaccacg ctcaccggct ccggatttat
     6421 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     6481 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     6541 gtttgcgcaa cgttgttgcc atcgctacag gcatcgtggt gtcacgctcg tcgtttggta
     6601 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     6661 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     6721 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     6781 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     6841 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     6901 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     6961 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     7021 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     7081 taagggcgac acggaaatgt tgaatactca tattcttcct ttttcaatat tattgaagca
     7141 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     7201 aaataggggt cagtgttaca accaattaac caattctgaa cattatcgcg agcccattta
     7261 tacctgaata tggctcataa caccccttgc tcatgaccaa aatcccttaa cgtgagttac
     7321 gcgcgcgtcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     7381 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     7441 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     7501 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttagcccacc acttcaagaa
     7561 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     7621 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     7681 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     7741 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     7801 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     7861 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     7921 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     7981 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     8041 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     8101 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaaggcgaga gtagggaact
     8161 gccaggcatc aaactaagca gaaggcccct gacggatggc ctttttgcgt ttctacaaac
     8221 tctttctgtg ttgtaaaacg acggccagtc ttaagctcgg gccccctggg cggttctgat
     8281 aacgagtaat cgttaatccg caaataacgt aaaaacccgc ttcggcgggt ttttttatgg
     8341 ggggagttta gggaaagagc atttgtcaga atatttaagg gcgcctgtca ctttgcttga
     8401 tatatgagaa ttatttaacc ttataaatga gaaaaaagca acgcacttta aataagatac
     8461 gttgcttttt cgattgatga acacctataa ttaaactatt catctattat ttatgatttt
     8521 ttgtatatac aatatttcta gtttgttaaa gagaattaag aaaataaatc tcgaaaataa
     8581 taaagggaaa atcagttttt gatatcaaaa ttatacatgt caacgataat acaaaatata
     8641 atacaaacta taagatgtta tcagtattta ttatcattta gaataaattt tgtgtcgccc
     8701 ttaattgtga gcggataaca attacgagct tcatgcacag tggcgttgac tcaatattgg
     8761 ccattagcca tattattcat tggttatata gcataaatca atattggcta ttggccattg
     8821 catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc aatatgaccg
     8881 ccatgtt
//
