LOCUS       sequence-303973-        8512 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 8512)
  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 8512)
  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-ICAM1-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8512
                     /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             4117..4152
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             4162..4185
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             5845..5889
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5890..5916
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(6056..6073)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6062..6173
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6291..6311)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6296..6625
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6476..6611
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6538..6557
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6673..6720
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6739..7338
                     /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(6739..6758)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7235..7255
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7468..7589
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7505..7524)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7559..7578
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7642..8433,8434..8502))
                     /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     8265..8284
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8503..8512,1..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 cggccgccca aacatctgtg
     2761 tccccaagca aagtgatcct gcctagaggc ggcagcgtgc tggtcacatg tagcacctct
     2821 tgcgaccagc ctaagctgct gggcatcgag acacccctgc ctaagaaaga gctgctgctg
     2881 cccggcaaca accggaaggt gtacgagctg agcaacgtgc aagaggacag ccagcctatg
     2941 tgctacagca actgccctga tggccagtct accgccaaga ccttcctgac cgtgtactgg
     3001 acccctgaga gagtggaact ggcccctctg ccttcttggc agcctgtggg caagaatctg
     3061 accctgagat gccaggtgga aggcggagcc cctagagcca atctgacagt ggttctgctg
     3121 cggggcgaga aagaactgaa gagggaacct gccgtgggcg agcctgctga agtgacaaca
     3181 acagtgctcg tgcggagaga tcaccacggc gccaatttca gctgccggac agaactggat
     3241 ctgaggccac agggcctcga gctgttcgag aataccagcg ctccttacca gctgcagacc
     3301 ttcgtgctgc ctgctacacc tcctcagctg gtgtctccca gagtgctgga agtggatacc
     3361 cagggcacag tcgtgtgcag cctggatggc ctgtttcctg tgtctgaggc ccaagtgcat
     3421 ctggccctgg gagatcagag actgaacccc acagtgacct acggcaacga cagcttttct
     3481 gccaaggcca gcgtgtccgt gacagccgaa gatgagggaa cccagaggct gacctgtgcc
     3541 gtgatcctgg gcaaccagag ccaagagaca ctgcagaccg tgaccatcta cagcttcccc
     3601 gctcctaatg tgatcctgac caagcctgag gtgtccgagg gcaccgaagt gacagtgaag
     3661 tgcgaggccc atcctcgggc caaagtgaca ctgaatggcg tgccagctca gccactggga
     3721 cctagagcac agctcctgct gaaagccaca cctgaggaca acggcagaag cttcagctgc
     3781 agcgccacac tggaagtcgc cggacagctg atccacaaga accagaccag agaactgaga
     3841 gtgctgtacg gccccagact ggacgagaga gattgccccg gaaattggac ctggcctgag
     3901 aacagccagc agacccctat gtgtcaggcc tggggaaatc ctctgcctga gctgaagtgc
     3961 ctgaaggacg gcacctttcc tctgccaatc ggcgagtctg tgaccgtgac aagagatctg
     4021 gaaggcacct acctgtgcag agccagatcc acacagggcg aagtgacccg gaaagtgacc
     4081 gtcaatgtgc tgagccccag atacgaaggc gcgccggaag atcaagttga tccaagattg
     4141 attgatggca agggcggatc tctggaggtg ctgttccagg gaccaggcgg aagcggcaat
     4201 cccggcggcg atctggcccc ccagatgctg cgcgagctgc aggagaccaa tgccgccctg
     4261 caggacgtgc gcgagctgct gcgccaccgc gtgaaggaga tcaccttcct gaagaacacc
     4321 gtgatggagt gcgacgcctg cggtatgcag ccctctagag gctccggctc catcatccca
     4381 gttgaggagg agaacccgga cttctggaac cgcgaggcag ccgaggccct gggtgccgcc
     4441 aagaagctgc agcctgcaca gacagccgcc aagaacctca tcatcttcct gggcgatggg
     4501 atgggggtgt ctacggtgac agctgccagg atcctaaaag ggcagaagaa ggacaaactg
     4561 gggcctgaga tacccctggc catggaccgc ttcccatatg tggctctgtc caagacatac
     4621 aatgtagaca aacatgtgcc agacagtgga gccacagcca cggcctacct gtgcggggtc
     4681 aagggcaact tccagaccat tggcttgagt gcagccgccc gctttaacca gtgcaacacg
     4741 acacgcggca acgaggtcat ctccgtgatg aatcgggcca agaaagcagg gaagtcagtg
     4801 ggagtggtaa ccaccacacg agtgcagcac gcctcgccag ccggcaccta cgcccacacg
     4861 gtgaaccgca actggtactc ggacgccgac gtgcctgcct cggcccgcca ggaggggtgc
     4921 caggacatcg ctacgcagct catctccaac atggacattg acgtgatcct aggtggaggc
     4981 cgaaagtaca tgtttcgcat gggaacccca gaccctgagt acccagatga ctacagccaa
     5041 ggtgggacca ggctggacgg gaagaatctg gtgcaggaat ggctggcgaa gcgccagggt
     5101 gcccggtatg tgtggaaccg cactgagctc atgcaggctt ccctggaccc gtctgtgacc
     5161 catctcatgg gtctctttga gcctggagac atgaaatacg agatccaccg agactccaca
     5221 ctggacccct ccctgatgga gatgacagag gctgccctgc gcctgctgag caggaacccc
     5281 cgcggcttct tcctcttcgt ggagggtggt cgcatcgacc atggtcatca tgaaagcagg
     5341 gcttaccggg cactgactga gacgatcatg ttcgacgacg ccattgagag ggcgggccag
     5401 ctcaccagcg aggaggacac gctgagcctc gtcactgccg accactccca cgtcttctcc
     5461 ttcggaggct accccctgcg agggagctcc atcttcgggc tggcccctgg caaggcccgg
     5521 gacaggaagg cctacacggt cctcctatac ggaaacggtc caggctatgt gctcaaggac
     5581 ggcgcccggc cggatgttac cgagagcgag agcgggagcc ccgagtatcg gcagcagtca
     5641 gcagtgcccc tggacgaaga gacccacgca ggcgaggacg tggcggtgtt cgcgcgcggc
     5701 ccgcaggcgc acctggttca cggcgtgcag gagcagacct tcatagcgca cgtcatggcc
     5761 ttcgccgcct gcctggagcc ctacaccgcc tgcgacctgg cgccccccgc cggcaccacc
     5821 gacgccgcgc acccgggtgg cggaggactg aacgacatct tcgaggctca gaaaatcgaa
     5881 tggcacgaac atcaccatca ccatcaccat caccattaag gccggcctgt agtcactggc
     5941 acaacgcgtt acccggtaag ccaatcgggt atacacggtc gtcatactgc agacagggtt
     6001 cttctacttt gcaagatagt cttgagtagt aaaataaata gatagagaaa atcagcctcg
     6061 actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc
     6121 ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc atcacaacac
     6181 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
     6241 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg
     6301 tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     6361 tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     6421 gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc
     6481 gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat
     6541 ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt gaggaggctt
     6601 ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca ttttcggatc
     6661 tgatcagcac gtgttgacaa ttaatcatcg gcatagtata tcggcatagt ataatacgac
     6721 aaggtgagga actaaaccat gaccgagtac aagcccacgg tgcgcctcgc cacccgcgac
     6781 gacgtcccca gggccgtacg caccctcgcc gccgcgttcg ccgactaccc cgccacgcgc
     6841 cacaccgtcg atccggaccg ccacatcgag cgggtcaccg agctgcaaga actcttcctc
     6901 acgcgcgtcg ggctcgacat cggcaaggtg tgggtcgcgg acgacggcgc cgcggtggcg
     6961 gtctggacca cgccggagag cgtcgaagcg ggggcggtgt tcgccgagat cggcccgcgc
     7021 atggccgagt tgagcggttc ccggctggcc gcgcagcaac agatggaagg cctcctggcg
     7081 ccgcaccggc ccaaggagcc cgcgtggttc ctggccaccg tcggcgtctc gcccgaccac
     7141 cagggcaagg gtctgggcag cgccgtcgtg ctccccggag tggaggcggc cgagcgcgcc
     7201 ggggtgcccg ccttcctgga gacatccgcg ccccgcaacc tccccttcta cgagcggctc
     7261 ggcttcaccg tcaccgccga cgtcgaggtg cccgaaggac cgcgcacctg gtgcatgacc
     7321 cgcaagcccg gtgcctgaca cgtgctacga gatttcgatt ccaccgccgc cttctatgaa
     7381 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat
     7441 ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa
     7501 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     7561 ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag
     7621 agcttggcgt aatcatggtc attaccaatg cttaatcagt gaggcaccta tctcagcgat
     7681 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     7741 ggagggctta ccatctggcc ccagcgctgc gatgataccg cgagaaccac gctcaccggc
     7801 tccggattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     7861 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     7921 gccagttaat agtttgcgca acgttgttgc catcgctaca ggcatcgtgg tgtcacgctc
     7981 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     8041 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     8101 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     8161 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     8221 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     8281 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     8341 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     8401 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     8461 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atattcttcc tt
//
