LOCUS       sequence-304057-        8776 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 8776)
  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 8776)
  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-LRRC4B-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8776
                     /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             4381..4416
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             4426..4449
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             6109..6153
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6154..6180
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(6320..6337)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6326..6437
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6555..6575)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6560..6889
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6740..6875
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6802..6821
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6937..6984
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7003..7602
                     /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(7003..7022)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7499..7519
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7732..7853
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7769..7788)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7823..7842
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7906..8697,8698..8766))
                     /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     8529..8548
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8767..8776,1..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 cggccgccgc tggcggaggc
     2761 ggagttgctg ttacatctgc tgcaggcgga ggaagccctc ctgctacatc ttgtcctgtg
     2821 gcctgcagct gtagcaacca ggccagcaga gtgatctgca ctagaagaga tctggccgag
     2881 gtgcccgcca gcattcccgt gaataccaga tacctgaacc tgcaagagaa cggcatccaa
     2941 gtgatccgga ccgacacctt caagcacctg agacacctgg aaatcctgca gctcagcaag
     3001 aacctcgtgc ggaagatcga agtgggcgcc tttaatggcc tgcctagcct gaacaccctg
     3061 gaactgttcg acaacagact gaccaccgtg cctacacagg ccttcgagta cctgagcaag
     3121 ctgagagagc tgtggctgcg gaacaacccc atcgagtcta tccccagcta cgccttcaat
     3181 agagtgccca gcctgagaag gctggacctg ggagaactga agcggctcga gtatatcagc
     3241 gaggccgcct ttgagggcct cgtgaacctg agatatctga atctgggcat gtgcaacctg
     3301 aaggacatcc ccaatctgac cgctctcgtg cggctggaag aactggaact gagcggcaac
     3361 aggctggatc tgattagacc tggcagcttc cagggcctga cctctctgag aaaactgtgg
     3421 ctcatgcacg cccaggtggc cacaatcgag agaaacgcct tcgacgacct gaagtccctc
     3481 gaggaactga atctgagcca caacaacctg atgagcctgc ctcacgacct gttcacccct
     3541 ctgcacagac tggaacgggt gcacctgaac cacaatcctt ggcactgcaa ctgcgacgtg
     3601 ctgtggctct cctggtggct gaaagaaacc gtgccaagca acaccacctg ttgcgccaga
     3661 tgtcatgccc ctgctggact gaagggcaga tatatcggcg agctggacca gagccacttc
     3721 acctgttatg cccctgtgat cgtggaacct cctaccgacc tgaacgtgac agaaggcatg
     3781 gccgccgagc tgaagtgtag aacaggcacc agcatgacca gcgtgaactg gctgacaccc
     3841 aatggcacac tgatgaccca cggcagctac agagtgcgga tctctgtgct gcacgacggc
     3901 accctgaact tcaccaatgt gaccgtgcag gacaccggcc agtacacctg tatggtcacc
     3961 aatagcgccg gcaataccac agcctctgcc acactgaatg tgtccgccgt tgatcctgtt
     4021 gctgctggcg gaacaggatc tggtggcgga ggacctggtg gatctggcgg tgttggaggt
     4081 ggaagcggcg gctacaccta ctttaccacc gtgaccgtgg aaaccctgga aacacagcct
     4141 ggcgaagaag ccctgcagcc tagaggaaca gagaaagagc ctcctggacc taccacagat
     4201 ggtgtttggg gcggaggtag acctggggat gctgcaggac ctgcctctag ctctacaaca
     4261 gcccctgctc ctagaagcag cagacccacc gagaaggcct ttacagtgcc tatcaccgac
     4321 gtgaccgaga acgccctgaa ggatctggac gacgtgatga agaccaccaa aggcgcgccg
     4381 gaagatcaag ttgatccaag attgattgat ggcaagggcg gatctctgga ggtgctgttc
     4441 cagggaccag gcggaagcgg caatcccggc ggcgatctgg ccccccagat gctgcgcgag
     4501 ctgcaggaga ccaatgccgc cctgcaggac gtgcgcgagc tgctgcgcca ccgcgtgaag
     4561 gagatcacct tcctgaagaa caccgtgatg gagtgcgacg cctgcggtat gcagccctct
     4621 agaggctccg gctccatcat cccagttgag gaggagaacc cggacttctg gaaccgcgag
     4681 gcagccgagg ccctgggtgc cgccaagaag ctgcagcctg cacagacagc cgccaagaac
     4741 ctcatcatct tcctgggcga tgggatgggg gtgtctacgg tgacagctgc caggatccta
     4801 aaagggcaga agaaggacaa actggggcct gagatacccc tggccatgga ccgcttccca
     4861 tatgtggctc tgtccaagac atacaatgta gacaaacatg tgccagacag tggagccaca
     4921 gccacggcct acctgtgcgg ggtcaagggc aacttccaga ccattggctt gagtgcagcc
     4981 gcccgcttta accagtgcaa cacgacacgc ggcaacgagg tcatctccgt gatgaatcgg
     5041 gccaagaaag cagggaagtc agtgggagtg gtaaccacca cacgagtgca gcacgcctcg
     5101 ccagccggca cctacgccca cacggtgaac cgcaactggt actcggacgc cgacgtgcct
     5161 gcctcggccc gccaggaggg gtgccaggac atcgctacgc agctcatctc caacatggac
     5221 attgacgtga tcctaggtgg aggccgaaag tacatgtttc gcatgggaac cccagaccct
     5281 gagtacccag atgactacag ccaaggtggg accaggctgg acgggaagaa tctggtgcag
     5341 gaatggctgg cgaagcgcca gggtgcccgg tatgtgtgga accgcactga gctcatgcag
     5401 gcttccctgg acccgtctgt gacccatctc atgggtctct ttgagcctgg agacatgaaa
     5461 tacgagatcc accgagactc cacactggac ccctccctga tggagatgac agaggctgcc
     5521 ctgcgcctgc tgagcaggaa cccccgcggc ttcttcctct tcgtggaggg tggtcgcatc
     5581 gaccatggtc atcatgaaag cagggcttac cgggcactga ctgagacgat catgttcgac
     5641 gacgccattg agagggcggg ccagctcacc agcgaggagg acacgctgag cctcgtcact
     5701 gccgaccact cccacgtctt ctccttcgga ggctaccccc tgcgagggag ctccatcttc
     5761 gggctggccc ctggcaaggc ccgggacagg aaggcctaca cggtcctcct atacggaaac
     5821 ggtccaggct atgtgctcaa ggacggcgcc cggccggatg ttaccgagag cgagagcggg
     5881 agccccgagt atcggcagca gtcagcagtg cccctggacg aagagaccca cgcaggcgag
     5941 gacgtggcgg tgttcgcgcg cggcccgcag gcgcacctgg ttcacggcgt gcaggagcag
     6001 accttcatag cgcacgtcat ggccttcgcc gcctgcctgg agccctacac cgcctgcgac
     6061 ctggcgcccc ccgccggcac caccgacgcc gcgcacccgg gtggcggagg actgaacgac
     6121 atcttcgagg ctcagaaaat cgaatggcac gaacatcacc atcaccatca ccatcaccat
     6181 taaggccggc ctgtagtcac tggcacaacg cgttacccgg taagccaatc gggtatacac
     6241 ggtcgtcata ctgcagacag ggttcttcta ctttgcaaga tagtcttgag tagtaaaata
     6301 aatagataga gaaaatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc
     6361 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa
     6421 aatgaggaaa ttgcatcaca acactcaacc ctatctcggt ctattctttt gatttataag
     6481 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     6541 cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     6601 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     6661 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     6721 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     6781 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct
     6841 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg
     6901 agcttgtata tccattttcg gatctgatca gcacgtgttg acaattaatc atcggcatag
     6961 tatatcggca tagtataata cgacaaggtg aggaactaaa ccatgaccga gtacaagccc
     7021 acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct cgccgccgcg
     7081 ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat cgagcgggtc
     7141 accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc
     7201 gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga agcgggggcg
     7261 gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct ggccgcgcag
     7321 caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc
     7381 accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc
     7441 ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacatc cgcgccccgc
     7501 aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa
     7561 ggaccgcgca cctggtgcat gacccgcaag cccggtgcct gacacgtgct acgagatttc
     7621 gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc
     7681 tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt
     7741 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
     7801 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc
     7861 tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcattacc aatgcttaat
     7921 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     7981 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagcg ctgcgatgat
     8041 accgcgagaa ccacgctcac cggctccgga tttatcagca ataaaccagc cagccggaag
     8101 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     8161 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccatcgc
     8221 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     8281 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     8341 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     8401 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     8461 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     8521 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     8581 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     8641 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     8701 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     8761 actcatattc ttcctt
//
