LOCUS       sequence-424379-        7705 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 7705)
  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 7705)
  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-FCGR2A-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..7705
                     /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             1544..1579
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             1589..1612
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             3272..3316
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             3317..3343
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(3483..3500)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3489..3600
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3718..3738)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3723..4052
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3903..4038
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3965..3984
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4100..4147
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4166..4765
                     /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(4166..4185)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4662..4682
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4895..5016
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4932..4951)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4986..5005
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(5069..5860,5861..5929))
                     /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     5692..5711
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5930..6021)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6063..6082
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6088..6107
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      6193..6781
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6682..6701
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6935..6952
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     7050..7067
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7051..7067
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    7523..7539
                     /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 cccaagctgc tgctcctcct aaagctgtgc tgaagctgga
     1021 acctccttgg atcaacgtgc tgcaagagga cagcgtgacc ctgacttgtc agggcgccag
     1081 atctcctgag agcgacagca tccagtggtt ccacaacggc aatctgatcc ccacacacac
     1141 ccagcctagc taccggttca aggccaacaa caacgacagc ggcgagtaca cctgtcagac
     1201 aggccagaca tctctgagcg atcctgtgca cctgacagtg ctgtctgaat ggctggtgct
     1261 gcagacccct cacctggaat ttcaagaggg cgagacaatc atgctgcggt gccacagctg
     1321 gaaggacaag cctctggtca aagtgacatt cttccagaac gggaagtccc agaagttcag
     1381 ccatctggac cccaccttta gcatccctca ggccaatcac agccactccg gcgattacca
     1441 ctgtaccggc aatatcggct acaccctgtt cagcagcaag cccgtgacaa tcaccgtgca
     1501 ggtcccatct atgggcagca gctctcctat gggaggcgcg ccggaagatc aagttgatcc
     1561 aagattgatt gatggcaagg gcggatctct ggaggtgctg ttccagggac caggcggaag
     1621 cggcaatccc ggcggcgatc tggcccccca gatgctgcgc gagctgcagg agaccaatgc
     1681 cgccctgcag gacgtgcgcg agctgctgcg ccaccgcgtg aaggagatca ccttcctgaa
     1741 gaacaccgtg atggagtgcg acgcctgcgg tatgcagccc tctagaggct ccggctccat
     1801 catcccagtt gaggaggaga acccggactt ctggaaccgc gaggcagccg aggccctggg
     1861 tgccgccaag aagctgcagc ctgcacagac agccgccaag aacctcatca tcttcctggg
     1921 cgatgggatg ggggtgtcta cggtgacagc tgccaggatc ctaaaagggc agaagaagga
     1981 caaactgggg cctgagatac ccctggccat ggaccgcttc ccatatgtgg ctctgtccaa
     2041 gacatacaat gtagacaaac atgtgccaga cagtggagcc acagccacgg cctacctgtg
     2101 cggggtcaag ggcaacttcc agaccattgg cttgagtgca gccgcccgct ttaaccagtg
     2161 caacacgaca cgcggcaacg aggtcatctc cgtgatgaat cgggccaaga aagcagggaa
     2221 gtcagtggga gtggtaacca ccacacgagt gcagcacgcc tcgccagccg gcacctacgc
     2281 ccacacggtg aaccgcaact ggtactcgga cgccgacgtg cctgcctcgg cccgccagga
     2341 ggggtgccag gacatcgcta cgcagctcat ctccaacatg gacattgacg tgatcctagg
     2401 tggaggccga aagtacatgt ttcgcatggg aaccccagac cctgagtacc cagatgacta
     2461 cagccaaggt gggaccaggc tggacgggaa gaatctggtg caggaatggc tggcgaagcg
     2521 ccagggtgcc cggtatgtgt ggaaccgcac tgagctcatg caggcttccc tggacccgtc
     2581 tgtgacccat ctcatgggtc tctttgagcc tggagacatg aaatacgaga tccaccgaga
     2641 ctccacactg gacccctccc tgatggagat gacagaggct gccctgcgcc tgctgagcag
     2701 gaacccccgc ggcttcttcc tcttcgtgga gggtggtcgc atcgaccatg gtcatcatga
     2761 aagcagggct taccgggcac tgactgagac gatcatgttc gacgacgcca ttgagagggc
     2821 gggccagctc accagcgagg aggacacgct gagcctcgtc actgccgacc actcccacgt
     2881 cttctccttc ggaggctacc ccctgcgagg gagctccatc ttcgggctgg cccctggcaa
     2941 ggcccgggac aggaaggcct acacggtcct cctatacgga aacggtccag gctatgtgct
     3001 caaggacggc gcccggccgg atgttaccga gagcgagagc gggagccccg agtatcggca
     3061 gcagtcagca gtgcccctgg acgaagagac ccacgcaggc gaggacgtgg cggtgttcgc
     3121 gcgcggcccg caggcgcacc tggttcacgg cgtgcaggag cagaccttca tagcgcacgt
     3181 catggccttc gccgcctgcc tggagcccta caccgcctgc gacctggcgc cccccgccgg
     3241 caccaccgac gccgcgcacc cgggtggcgg aggactgaac gacatcttcg aggctcagaa
     3301 aatcgaatgg cacgaacatc accatcacca tcaccatcac cattaaggcc ggcctgtagt
     3361 cactggcaca acgcgttacc cggtaagcca atcgggtata cacggtcgtc atactgcaga
     3421 cagggttctt ctactttgca agatagtctt gagtagtaaa ataaatagat agagaaaatc
     3481 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     3541 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     3601 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     3661 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga
     3721 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     3781 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
     3841 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     3901 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     3961 tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag
     4021 gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt
     4081 tcggatctga tcagcacgtg ttgacaatta atcatcggca tagtatatcg gcatagtata
     4141 atacgacaag gtgaggaact aaaccatgac cgagtacaag cccacggtgc gcctcgccac
     4201 ccgcgacgac gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc
     4261 cacgcgccac accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact
     4321 cttcctcacg cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc
     4381 ggtggcggtc tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg
     4441 cccgcgcatg gccgagttga gcggttcccg gctggccgcg cagcaacaga tggaaggcct
     4501 cctggcgccg caccggccca aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc
     4561 cgaccaccag ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga
     4621 gcgcgccggg gtgcccgcct tcctggagac atccgcgccc cgcaacctcc ccttctacga
     4681 gcggctcggc ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg
     4741 catgacccgc aagcccggtg cctgacacgt gctacgagat ttcgattcca ccgccgcctt
     4801 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg
     4861 cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg
     4921 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     4981 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc
     5041 tagctagagc ttggcgtaat catggtcatt accaatgctt aatcagtgag gcacctatct
     5101 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     5161 cgatacggga gggcttacca tctggcccca gcgctgcgat gataccgcga gaaccacgct
     5221 caccggctcc ggatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     5281 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     5341 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat cgctacaggc atcgtggtgt
     5401 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     5461 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     5521 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     5581 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     5641 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     5701 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     5761 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     5821 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     5881 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ttcttccttt
     5941 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     6001 gtatttagaa aaataaacaa ataggggtca gtgttacaac caattaacca attctgaaca
     6061 ttatcgcgag cccatttata cctgaatatg gctcataaca ccccttgctc atgaccaaaa
     6121 tcccttaacg tgagttacgc gcgcgtcgtt ccactgagcg tcagaccccg tagaaaagat
     6181 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     6241 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     6301 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt
     6361 agcccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     6421 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     6481 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     6541 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     6601 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     6661 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     6721 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     6781 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     6841 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc
     6901 tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga
     6961 aggcgagagt agggaactgc caggcatcaa actaagcaga aggcccctga cggatggcct
     7021 ttttgcgttt ctacaaactc tttctgtgtt gtaaaacgac ggccagtctt aagctcgggc
     7081 cccctgggcg gttctgataa cgagtaatcg ttaatccgca aataacgtaa aaacccgctt
     7141 cggcgggttt ttttatgggg ggagtttagg gaaagagcat ttgtcagaat atttaagggc
     7201 gcctgtcact ttgcttgata tatgagaatt atttaacctt ataaatgaga aaaaagcaac
     7261 gcactttaaa taagatacgt tgctttttcg attgatgaac acctataatt aaactattca
     7321 tctattattt atgatttttt gtatatacaa tatttctagt ttgttaaaga gaattaagaa
     7381 aataaatctc gaaaataata aagggaaaat cagtttttga tatcaaaatt atacatgtca
     7441 acgataatac aaaatataat acaaactata agatgttatc agtatttatt atcatttaga
     7501 ataaattttg tgtcgccctt aattgtgagc ggataacaat tacgagcttc atgcacagtg
     7561 gcgttgactc aatattggcc attagccata ttattcattg gttatatagc ataaatcaat
     7621 attggctatt ggccattgca tacgttgtat ctatatcata atatgtacat ttatattggc
     7681 tcatgtccaa tatgaccgcc atgtt
//
