LOCUS       sequence-303793-        8170 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 8170)
  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 8170)
  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-IL1RAPL1-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8170
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     124..143
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     149..168
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      254..842
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     743..762
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     996..1013
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     1111..1128
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1112..1128
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1584..1600
                     /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        1767..2146
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2147..2350
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2301..2321
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2347..2371
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        2581..2599
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2581..2598
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     2612..2631
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             3775..3810
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             3820..3843
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             5503..5547
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5548..5574
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(5714..5731)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5720..5831
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5949..5969)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5954..6283
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6134..6269
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6196..6215
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6331..6378
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6397..6996
                     /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(6397..6416)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6893..6913
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7126..7247
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7163..7182)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7217..7236
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7300..8091,8092..8160))
                     /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     7923..7942
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8161..8170,1..82))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
       61 tgtatttaga aaaataaaca aataggggtc agtgttacaa ccaattaacc aattctgaac
      121 attatcgcga gcccatttat acctgaatat ggctcataac accccttgct catgaccaaa
      181 atcccttaac gtgagttacg cgcgcgtcgt tccactgagc gtcagacccc gtagaaaaga
      241 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
      301 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
      361 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
      421 tagcccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
      481 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
      541 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
      601 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
      661 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
      721 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
      781 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
      841 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
      901 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag
      961 ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg
     1021 aaggcgagag tagggaactg ccaggcatca aactaagcag aaggcccctg acggatggcc
     1081 tttttgcgtt tctacaaact ctttctgtgt tgtaaaacga cggccagtct taagctcggg
     1141 ccccctgggc ggttctgata acgagtaatc gttaatccgc aaataacgta aaaacccgct
     1201 tcggcgggtt tttttatggg gggagtttag ggaaagagca tttgtcagaa tatttaaggg
     1261 cgcctgtcac tttgcttgat atatgagaat tatttaacct tataaatgag aaaaaagcaa
     1321 cgcactttaa ataagatacg ttgctttttc gattgatgaa cacctataat taaactattc
     1381 atctattatt tatgattttt tgtatataca atatttctag tttgttaaag agaattaaga
     1441 aaataaatct cgaaaataat aaagggaaaa tcagtttttg atatcaaaat tatacatgtc
     1501 aacgataata caaaatataa tacaaactat aagatgttat cagtatttat tatcatttag
     1561 aataaatttt gtgtcgccct taattgtgag cggataacaa ttacgagctt catgcacagt
     1621 ggcgttgact caatattggc cattagccat attattcatt ggttatatag cataaatcaa
     1681 tattggctat tggccattgc atacgttgta tctatatcat aatatgtaca tttatattgg
     1741 ctcatgtcca atatgaccgc catgttggca ttgattattg actagttatt aatagtaatc
     1801 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
     1861 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
     1921 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
     1981 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
     2041 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
     2101 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
     2161 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
     2221 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
     2281 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
     2341 aagcagagct cgtttagtga accgtcagat cgcctggaga cgccatccac gctgttttga
     2401 cctccataga agacaccggg accgatccag cctccgcggc cgggaacggt gcattggaac
     2461 gcggattccc cgtgccaaga gtgacgtaag taccgcctat agagtctata ggcccacccc
     2521 cttggcttcg ttagaacgcg gctacaatta atacataacc ttatgtatca tacacatacg
     2581 atttaggtga cactatagaa taacatccac tttgcctttc tctccacagg tgtccactcc
     2641 caggtccaac tgcacctcgg ttctatcgaa aacgcgcctg ctagcgccac catgaagacc
     2701 atcattgccc tgagctacat cttctgcctg gtgttcgctg cggccgcctt gaaggttgtg
     2761 acaaagagag gaagcgccga cggctgcacc gattggagca tcgacatcaa gaaataccag
     2821 gtgctcgtgg gcgagcccgt gcggattaag tgtgccctgt tttacggcta catccggacc
     2881 aactacagcc tggctcagtc tgccggactg tccctgatgt ggtataagtc tagcggccct
     2941 ggcgacttcg aggaacctat cgcctttgac ggcagccgga tgagcaaaga agaggacagc
     3001 atctggttca gacccacact gctgcaggat agcggcctgt acgcctgcgt gatcagaaac
     3061 agcacctact gcatgaaggt gtccatcagc ctgacagtgg gcgagaatga taccggcctg
     3121 tgctacaaca gcaagatgaa gtactttgag aaggccgagc tgagcaagag caaagagatc
     3181 agctgccggg acatcgagga ctttctgctg cctaccagag aacccgagat cctgtggtac
     3241 aaagagtgca ggaccaagac ctggcggccc agcatcgtgt tcaagagaga caccctgctg
     3301 atccgggaag tgcgggaaga tgacatcggc aactacacat gcgagctgaa gtacggcggc
     3361 ttcgtcgtgc ggagaacaac cgaactgaca gtgacagccc ctctgaccga caagcctcct
     3421 aagctgctgt accccatgga aagcaagctg accatccaag agacacagct gggcgacagc
     3481 gccaatctga cctgtagagc cttttttggc tactccggcg acgtgtcccc tctgatctat
     3541 tggatgaagg gcgagaagtt catcgaggat ctggacgaga acagagtgtg ggagagcgac
     3601 atcagaatcc tgaaagaaca cctgggcgag caagaagtgt ccatctctct gatcgtggac
     3661 agcgtggaag agggcgacct gggcaattac agctgctacg tggaaaacgg caatggcaga
     3721 cggcacgcct ctgttctgct gcacaagcgg gaactgatgt acacaggcgc gccggaagat
     3781 caagttgatc caagattgat tgatggcaag ggcggatctc tggaggtgct gttccaggga
     3841 ccaggcggaa gcggcaatcc cggcggcgat ctggcccccc agatgctgcg cgagctgcag
     3901 gagaccaatg ccgccctgca ggacgtgcgc gagctgctgc gccaccgcgt gaaggagatc
     3961 accttcctga agaacaccgt gatggagtgc gacgcctgcg gtatgcagcc ctctagaggc
     4021 tccggctcca tcatcccagt tgaggaggag aacccggact tctggaaccg cgaggcagcc
     4081 gaggccctgg gtgccgccaa gaagctgcag cctgcacaga cagccgccaa gaacctcatc
     4141 atcttcctgg gcgatgggat gggggtgtct acggtgacag ctgccaggat cctaaaaggg
     4201 cagaagaagg acaaactggg gcctgagata cccctggcca tggaccgctt cccatatgtg
     4261 gctctgtcca agacatacaa tgtagacaaa catgtgccag acagtggagc cacagccacg
     4321 gcctacctgt gcggggtcaa gggcaacttc cagaccattg gcttgagtgc agccgcccgc
     4381 tttaaccagt gcaacacgac acgcggcaac gaggtcatct ccgtgatgaa tcgggccaag
     4441 aaagcaggga agtcagtggg agtggtaacc accacacgag tgcagcacgc ctcgccagcc
     4501 ggcacctacg cccacacggt gaaccgcaac tggtactcgg acgccgacgt gcctgcctcg
     4561 gcccgccagg aggggtgcca ggacatcgct acgcagctca tctccaacat ggacattgac
     4621 gtgatcctag gtggaggccg aaagtacatg tttcgcatgg gaaccccaga ccctgagtac
     4681 ccagatgact acagccaagg tgggaccagg ctggacggga agaatctggt gcaggaatgg
     4741 ctggcgaagc gccagggtgc ccggtatgtg tggaaccgca ctgagctcat gcaggcttcc
     4801 ctggacccgt ctgtgaccca tctcatgggt ctctttgagc ctggagacat gaaatacgag
     4861 atccaccgag actccacact ggacccctcc ctgatggaga tgacagaggc tgccctgcgc
     4921 ctgctgagca ggaacccccg cggcttcttc ctcttcgtgg agggtggtcg catcgaccat
     4981 ggtcatcatg aaagcagggc ttaccgggca ctgactgaga cgatcatgtt cgacgacgcc
     5041 attgagaggg cgggccagct caccagcgag gaggacacgc tgagcctcgt cactgccgac
     5101 cactcccacg tcttctcctt cggaggctac cccctgcgag ggagctccat cttcgggctg
     5161 gcccctggca aggcccggga caggaaggcc tacacggtcc tcctatacgg aaacggtcca
     5221 ggctatgtgc tcaaggacgg cgcccggccg gatgttaccg agagcgagag cgggagcccc
     5281 gagtatcggc agcagtcagc agtgcccctg gacgaagaga cccacgcagg cgaggacgtg
     5341 gcggtgttcg cgcgcggccc gcaggcgcac ctggttcacg gcgtgcagga gcagaccttc
     5401 atagcgcacg tcatggcctt cgccgcctgc ctggagccct acaccgcctg cgacctggcg
     5461 ccccccgccg gcaccaccga cgccgcgcac ccgggtggcg gaggactgaa cgacatcttc
     5521 gaggctcaga aaatcgaatg gcacgaacat caccatcacc atcaccatca ccattaaggc
     5581 cggcctgtag tcactggcac aacgcgttac ccggtaagcc aatcgggtat acacggtcgt
     5641 catactgcag acagggttct tctactttgc aagatagtct tgagtagtaa aataaataga
     5701 tagagaaaat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc
     5761 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag
     5821 gaaattgcat cacaacactc aaccctatct cggtctattc ttttgattta taagggattt
     5881 tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     5941 aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag
     6001 aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc
     6061 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc
     6121 cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg
     6181 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca
     6241 gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg
     6301 tatatccatt ttcggatctg atcagcacgt gttgacaatt aatcatcggc atagtatatc
     6361 ggcatagtat aatacgacaa ggtgaggaac taaaccatga ccgagtacaa gcccacggtg
     6421 cgcctcgcca cccgcgacga cgtccccagg gccgtacgca ccctcgccgc cgcgttcgcc
     6481 gactaccccg ccacgcgcca caccgtcgat ccggaccgcc acatcgagcg ggtcaccgag
     6541 ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac
     6601 gacggcgccg cggtggcggt ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc
     6661 gccgagatcg gcccgcgcat ggccgagttg agcggttccc ggctggccgc gcagcaacag
     6721 atggaaggcc tcctggcgcc gcaccggccc aaggagcccg cgtggttcct ggccaccgtc
     6781 ggcgtctcgc ccgaccacca gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg
     6841 gaggcggccg agcgcgccgg ggtgcccgcc ttcctggaga catccgcgcc ccgcaacctc
     6901 cccttctacg agcggctcgg cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg
     6961 cgcacctggt gcatgacccg caagcccggt gcctgacacg tgctacgaga tttcgattcc
     7021 accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg
     7081 atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca
     7141 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt
     7201 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata
     7261 ccgtcgacct ctagctagag cttggcgtaa tcatggtcat taccaatgct taatcagtga
     7321 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     7381 gtagataact acgatacggg agggcttacc atctggcccc agcgctgcga tgataccgcg
     7441 agaaccacgc tcaccggctc cggatttatc agcaataaac cagccagccg gaagggccga
     7501 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     7561 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca tcgctacagg
     7621 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     7681 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     7741 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     7801 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     7861 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     7921 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     7981 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     8041 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     8101 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     8161 attcttcctt
//
