LOCUS       sequence-474346-        8485 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 8485)
  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 8485)
  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-LILRA1-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8485
                     /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             4090..4125
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             4135..4158
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             5818..5862
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5863..5889
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(6029..6046)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6035..6146
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6264..6284)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6269..6598
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6449..6584
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6511..6530
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6646..6693
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6712..7311
                     /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(6712..6731)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7208..7228
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7441..7562
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7478..7497)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7532..7551
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7615..8406,8407..8475))
                     /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     8238..8257
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8476..8485,1..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 cggccgccag aacacatgtt
     2761 caggctggca cactgcccaa gcctacactg tgggctgaac ctggcagcgt gatcacacag
     2821 ggaagccctg tgacactgtg gtgccagggc atcctggaaa cccaagagta ccggctgtac
     2881 cgcgagaaga aaacagcccc ttggatcacc agaattcccc aagagatcgt gaagaagggg
     2941 cagttcccca ttcctagcat cacctgggag cacaccggca gatacagatg cttctacggc
     3001 tctcataccg ccggatggtc cgagccttct gaccctctgg aactggtggt cacaggcgcc
     3061 tacatcaagc ccactctgtc tgccctgcct tctccagtgg ttacctctgg cggaaacgtg
     3121 accctgcact gtgtgtctca ggtggccttc ggcagcttca tcctgtgcaa agaaggcgag
     3181 gacgagcacc ctcagtgcct gaatagccag ccaagaacac acggctggtc ccgggccatc
     3241 ttttctgtgg gacctgtgtc tcctagccgg cggtggtcct acagatgtta cgcctacgac
     3301 agcaacagcc ctcacgtgtg gtccctgcca agcgatctgc tggaactgct cgtgctgggc
     3361 gtgtccaaga aacctagcct gtctgtgcag cccggaccta ttgtggctcc tggcgaatct
     3421 ctgactctgc agtgcgtgtc cgacgtgtcc tacgatagat tcgtgctcta caaagagggc
     3481 gagagagact tcctgcagct gcctggacct cagcctcaag ctggactgag ccaggccaat
     3541 ttcacactgg gccctgtgtc cagaagctac ggcggccagt atagatgtag cggcgcctat
     3601 aacctgagca gcgagtggtc cgctcctagc gatcctctgg atatcctgat cgccggccag
     3661 ttcagaggca gaccctttat ctctgtgcac cctgggccta cagtggcctc tggcgagaat
     3721 gtgactctgc tgtgtcagag ctggggcccc ttccacacct ttctgctgac aaaagccggc
     3781 gctgccgacg ctcctctgag actgagatct atccacgagt accccaagta ccaggccgag
     3841 tttcctatga gcccagtgac atctgcccac agcggcacct atagatgcta cggcagcctg
     3901 tccagcaatc cctacctgct gtctcacccc agcgacagcc tggaactcat ggtttctggc
     3961 gccgcagaga cactgagccc tccacagaac aagagcgaca gcaaagcagg cgcagccaac
     4021 acactgtccc caagccagaa caaaaccgcc agccatcctc aggactacac cgtggaaaat
     4081 ggcgcgccgg aagatcaagt tgatccaaga ttgattgatg gcaagggcgg atctctggag
     4141 gtgctgttcc agggaccagg cggaagcggc aatcccggcg gcgatctggc cccccagatg
     4201 ctgcgcgagc tgcaggagac caatgccgcc ctgcaggacg tgcgcgagct gctgcgccac
     4261 cgcgtgaagg agatcacctt cctgaagaac accgtgatgg agtgcgacgc ctgcggtatg
     4321 cagccctcta gaggctccgg ctccatcatc ccagttgagg aggagaaccc ggacttctgg
     4381 aaccgcgagg cagccgaggc cctgggtgcc gccaagaagc tgcagcctgc acagacagcc
     4441 gccaagaacc tcatcatctt cctgggcgat gggatggggg tgtctacggt gacagctgcc
     4501 aggatcctaa aagggcagaa gaaggacaaa ctggggcctg agatacccct ggccatggac
     4561 cgcttcccat atgtggctct gtccaagaca tacaatgtag acaaacatgt gccagacagt
     4621 ggagccacag ccacggccta cctgtgcggg gtcaagggca acttccagac cattggcttg
     4681 agtgcagccg cccgctttaa ccagtgcaac acgacacgcg gcaacgaggt catctccgtg
     4741 atgaatcggg ccaagaaagc agggaagtca gtgggagtgg taaccaccac acgagtgcag
     4801 cacgcctcgc cagccggcac ctacgcccac acggtgaacc gcaactggta ctcggacgcc
     4861 gacgtgcctg cctcggcccg ccaggagggg tgccaggaca tcgctacgca gctcatctcc
     4921 aacatggaca ttgacgtgat cctaggtgga ggccgaaagt acatgtttcg catgggaacc
     4981 ccagaccctg agtacccaga tgactacagc caaggtggga ccaggctgga cgggaagaat
     5041 ctggtgcagg aatggctggc gaagcgccag ggtgcccggt atgtgtggaa ccgcactgag
     5101 ctcatgcagg cttccctgga cccgtctgtg acccatctca tgggtctctt tgagcctgga
     5161 gacatgaaat acgagatcca ccgagactcc acactggacc cctccctgat ggagatgaca
     5221 gaggctgccc tgcgcctgct gagcaggaac ccccgcggct tcttcctctt cgtggagggt
     5281 ggtcgcatcg accatggtca tcatgaaagc agggcttacc gggcactgac tgagacgatc
     5341 atgttcgacg acgccattga gagggcgggc cagctcacca gcgaggagga cacgctgagc
     5401 ctcgtcactg ccgaccactc ccacgtcttc tccttcggag gctaccccct gcgagggagc
     5461 tccatcttcg ggctggcccc tggcaaggcc cgggacagga aggcctacac ggtcctccta
     5521 tacggaaacg gtccaggcta tgtgctcaag gacggcgccc ggccggatgt taccgagagc
     5581 gagagcggga gccccgagta tcggcagcag tcagcagtgc ccctggacga agagacccac
     5641 gcaggcgagg acgtggcggt gttcgcgcgc ggcccgcagg cgcacctggt tcacggcgtg
     5701 caggagcaga ccttcatagc gcacgtcatg gccttcgccg cctgcctgga gccctacacc
     5761 gcctgcgacc tggcgccccc cgccggcacc accgacgccg cgcacccggg tggcggagga
     5821 ctgaacgaca tcttcgaggc tcagaaaatc gaatggcacg aacatcacca tcaccatcac
     5881 catcaccatt aaggccggcc tgtagtcact ggcacaacgc gttacccggt aagccaatcg
     5941 ggtatacacg gtcgtcatac tgcagacagg gttcttctac tttgcaagat agtcttgagt
     6001 agtaaaataa atagatagag aaaatcagcc tcgactgtgc cttctagttg ccagccatct
     6061 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt
     6121 tcctaataaa atgaggaaat tgcatcacaa cactcaaccc tatctcggtc tattcttttg
     6181 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     6241 aatttaacgc gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
     6301 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     6361 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     6421 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     6481 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc
     6541 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     6601 gctcccggga gcttgtatat ccattttcgg atctgatcag cacgtgttga caattaatca
     6661 tcggcatagt atatcggcat agtataatac gacaaggtga ggaactaaac catgaccgag
     6721 tacaagccca cggtgcgcct cgccacccgc gacgacgtcc ccagggccgt acgcaccctc
     6781 gccgccgcgt tcgccgacta ccccgccacg cgccacaccg tcgatccgga ccgccacatc
     6841 gagcgggtca ccgagctgca agaactcttc ctcacgcgcg tcgggctcga catcggcaag
     6901 gtgtgggtcg cggacgacgg cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa
     6961 gcgggggcgg tgttcgccga gatcggcccg cgcatggccg agttgagcgg ttcccggctg
     7021 gccgcgcagc aacagatgga aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg
     7081 ttcctggcca ccgtcggcgt ctcgcccgac caccagggca agggtctggg cagcgccgtc
     7141 gtgctccccg gagtggaggc ggccgagcgc gccggggtgc ccgccttcct ggagacatcc
     7201 gcgccccgca acctcccctt ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag
     7261 gtgcccgaag gaccgcgcac ctggtgcatg acccgcaagc ccggtgcctg acacgtgcta
     7321 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg
     7381 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc
     7441 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     7501 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     7561 tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcattacca
     7621 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     7681 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagcgc
     7741 tgcgatgata ccgcgagaac cacgctcacc ggctccggat ttatcagcaa taaaccagcc
     7801 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     7861 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     7921 tgccatcgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     7981 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     8041 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     8101 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     8161 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     8221 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     8281 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     8341 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     8401 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     8461 atgttgaata ctcatattct tcctt
//
