LOCUS       sequence-304097-        9028 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 9028)
  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 9028)
  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-SEMA4F-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..9028
                     /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             4633..4668
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             4678..4701
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             6361..6405
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6406..6432
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(6572..6589)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6578..6689
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6807..6827)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6812..7141
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6992..7127
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7054..7073
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7189..7236
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7255..7854
                     /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(7255..7274)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7751..7771
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7984..8105
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8021..8040)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8075..8094
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(8158..8949,8950..9018))
                     /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     8781..8800
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(9019..9028,1..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 agttcctaga
     2761 agcgtgccaa gaaccagcct gcctatcagc gaggccgata gctgcctgac cagatttgcc
     2821 gtgcctcaca cctacaacta cagcgtgctg ctggtggacc ctgccagcca cacactgtat
     2881 gtgggcgcca gagataccat cttcgccctg agcctgcctt ttagcggcga gcggcctaga
     2941 agaatcgact ggatggtgcc cgaggctcac cggcagaact gtagaaagaa gggcaagaaa
     3001 gaggacgagt gccacaactt cgtgcagatc ctggccattg ccaacgccag ccatctgctg
     3061 acctgtggca ccttcgcctt cgatcctaag tgcggcgtga tcgacgtgtc cagattccag
     3121 caggtcgagc ggctggaaag cggcagaggc aagtgtcctt ttgagcccgc tcagagatct
     3181 gccgctgtta tggcaggcgg agtgctgtat gccgccaccg tgaagaatta cctgggcacc
     3241 gagcctatca tcaccagagc tgttggcaga gccgaggact ggatcagaac cgacacactg
     3301 cccagctggc tgaatgcccc tgcttttgtg gctgccgtgg ctctgtctcc tgccgaatgg
     3361 ggagatgagg atggcgacga cgagatctac ttcttcttca ccgaaacctc cagagccttc
     3421 gacagctacg agcggatcaa ggtgccaaga gtggccagag tgtgtgccgg cgatctcggc
     3481 ggaagaaaga cactgcagca gcggtggacc accttcctga aagccgatct gctgtgccct
     3541 ggacctgaac acggtagagc tagttccgtg ctgcaggatg tggctgtgct gaggcctgaa
     3601 cttggagccg gaacacccat cttctacggc atcttcagca gccagtggga gggcgccaca
     3661 atctctgccg tgtgtgcctt cagaccccaa gacatcagga ccgtgctgaa cggccctttc
     3721 agagagctga agcacgactg caacagaggc ctgcctgtgg tggacaacga tgtgcctcag
     3781 cctagacctg gcgagtgcat caccaacaac atgaagctgc ggcacttcgg cagcagcctg
     3841 tctctgcctg atagagtgct gacctttatc cgggatcacc ctctgatgga ccggcctgtg
     3901 tttcccgctg atggacaccc tctgctggtc accacagata ccgcctacct gagagtggtg
     3961 gcccacagag tgacaagcct gagcggcaaa gaatacgacg tgctgtatct gggcacagag
     4021 gacggccatc tgcacagagc cgttagaatt ggcgcccagc tgagcgtgct ggaagatctg
     4081 gctctgttcc ctgagccaca gcctgtggaa aacatgaagt tgtaccatag ctggctgctc
     4141 gtgggcagca gaaccgaagt gacccaagtg aacaccacca actgcggcag actgcagagc
     4201 tgctccgagt gtatcctggc tcaggaccct gtgtgcgcct ggtcctttag actggatgaa
     4261 tgtgtggccc acgccggcga gcatagagga ctggtgcagg atattgagag cgccgatgtg
     4321 tccagcctgt gtcctaaaga gcccggcgaa agacccgtgg tgtttgaagt gcctgtggcc
     4381 actgctgctc acgtggtgtt gccttgttct cctagctctg cctgggccag ctgtgtgtgg
     4441 catcaaccta gcggagtgac cgctctgacc cctagaagag atggcctgga agtggtggtt
     4501 acccctggcg ccatgggagc ctatgcctgc gaatgtcaag aaggcggagc agcccacgtg
     4561 gtggccgctt attctcttgt gtggggcagc cagagagatg cccctagcag agcccatacc
     4621 gttggcgcgc cggaagatca agttgatcca agattgattg atggcaaggg cggatctctg
     4681 gaggtgctgt tccagggacc aggcggaagc ggcaatcccg gcggcgatct ggccccccag
     4741 atgctgcgcg agctgcagga gaccaatgcc gccctgcagg acgtgcgcga gctgctgcgc
     4801 caccgcgtga aggagatcac cttcctgaag aacaccgtga tggagtgcga cgcctgcggt
     4861 atgcagccct ctagaggctc cggctccatc atcccagttg aggaggagaa cccggacttc
     4921 tggaaccgcg aggcagccga ggccctgggt gccgccaaga agctgcagcc tgcacagaca
     4981 gccgccaaga acctcatcat cttcctgggc gatgggatgg gggtgtctac ggtgacagct
     5041 gccaggatcc taaaagggca gaagaaggac aaactggggc ctgagatacc cctggccatg
     5101 gaccgcttcc catatgtggc tctgtccaag acatacaatg tagacaaaca tgtgccagac
     5161 agtggagcca cagccacggc ctacctgtgc ggggtcaagg gcaacttcca gaccattggc
     5221 ttgagtgcag ccgcccgctt taaccagtgc aacacgacac gcggcaacga ggtcatctcc
     5281 gtgatgaatc gggccaagaa agcagggaag tcagtgggag tggtaaccac cacacgagtg
     5341 cagcacgcct cgccagccgg cacctacgcc cacacggtga accgcaactg gtactcggac
     5401 gccgacgtgc ctgcctcggc ccgccaggag gggtgccagg acatcgctac gcagctcatc
     5461 tccaacatgg acattgacgt gatcctaggt ggaggccgaa agtacatgtt tcgcatggga
     5521 accccagacc ctgagtaccc agatgactac agccaaggtg ggaccaggct ggacgggaag
     5581 aatctggtgc aggaatggct ggcgaagcgc cagggtgccc ggtatgtgtg gaaccgcact
     5641 gagctcatgc aggcttccct ggacccgtct gtgacccatc tcatgggtct ctttgagcct
     5701 ggagacatga aatacgagat ccaccgagac tccacactgg acccctccct gatggagatg
     5761 acagaggctg ccctgcgcct gctgagcagg aacccccgcg gcttcttcct cttcgtggag
     5821 ggtggtcgca tcgaccatgg tcatcatgaa agcagggctt accgggcact gactgagacg
     5881 atcatgttcg acgacgccat tgagagggcg ggccagctca ccagcgagga ggacacgctg
     5941 agcctcgtca ctgccgacca ctcccacgtc ttctccttcg gaggctaccc cctgcgaggg
     6001 agctccatct tcgggctggc ccctggcaag gcccgggaca ggaaggccta cacggtcctc
     6061 ctatacggaa acggtccagg ctatgtgctc aaggacggcg cccggccgga tgttaccgag
     6121 agcgagagcg ggagccccga gtatcggcag cagtcagcag tgcccctgga cgaagagacc
     6181 cacgcaggcg aggacgtggc ggtgttcgcg cgcggcccgc aggcgcacct ggttcacggc
     6241 gtgcaggagc agaccttcat agcgcacgtc atggccttcg ccgcctgcct ggagccctac
     6301 accgcctgcg acctggcgcc ccccgccggc accaccgacg ccgcgcaccc gggtggcgga
     6361 ggactgaacg acatcttcga ggctcagaaa atcgaatggc acgaacatca ccatcaccat
     6421 caccatcacc attaaggccg gcctgtagtc actggcacaa cgcgttaccc ggtaagccaa
     6481 tcgggtatac acggtcgtca tactgcagac agggttcttc tactttgcaa gatagtcttg
     6541 agtagtaaaa taaatagata gagaaaatca gcctcgactg tgccttctag ttgccagcca
     6601 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
     6661 ctttcctaat aaaatgagga aattgcatca caacactcaa ccctatctcg gtctattctt
     6721 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     6781 aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     6841 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     6901 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     6961 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     7021 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     7081 tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     7141 aaagctcccg ggagcttgta tatccatttt cggatctgat cagcacgtgt tgacaattaa
     7201 tcatcggcat agtatatcgg catagtataa tacgacaagg tgaggaacta aaccatgacc
     7261 gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg tccccagggc cgtacgcacc
     7321 ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca ccgtcgatcc ggaccgccac
     7381 atcgagcggg tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct cgacatcggc
     7441 aaggtgtggg tcgcggacga cggcgccgcg gtggcggtct ggaccacgcc ggagagcgtc
     7501 gaagcggggg cggtgttcgc cgagatcggc ccgcgcatgg ccgagttgag cggttcccgg
     7561 ctggccgcgc agcaacagat ggaaggcctc ctggcgccgc accggcccaa ggagcccgcg
     7621 tggttcctgg ccaccgtcgg cgtctcgccc gaccaccagg gcaagggtct gggcagcgcc
     7681 gtcgtgctcc ccggagtgga ggcggccgag cgcgccgggg tgcccgcctt cctggagaca
     7741 tccgcgcccc gcaacctccc cttctacgag cggctcggct tcaccgtcac cgccgacgtc
     7801 gaggtgcccg aaggaccgcg cacctggtgc atgacccgca agcccggtgc ctgacacgtg
     7861 ctacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc
     7921 cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac
     7981 cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     8041 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     8101 tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatta
     8161 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     8221 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     8281 cgctgcgatg ataccgcgag aaccacgctc accggctccg gatttatcag caataaacca
     8341 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     8401 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     8461 tgttgccatc gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     8521 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     8581 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     8641 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     8701 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     8761 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     8821 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     8881 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     8941 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     9001 gaaatgttga atactcatat tcttcctt
//
