LOCUS       sequence-400989-        8944 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 8944)
  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 8944)
  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-IL6ST-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8944
                     /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             2783..2818
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             2828..2851
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             4511..4555
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             4556..4582
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(4722..4739)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4728..4839
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4957..4977)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4962..5291
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5142..5277
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5204..5223
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        5339..5386
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5405..6004
                     /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(5405..5424)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5901..5921
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6134..6255
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6171..6190)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6225..6244
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(6308..7099,7100..7168))
                     /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     6931..6950
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7169..7260)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7302..7321
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     7327..7346
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      7432..8020
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7921..7940
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8174..8191
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     8289..8306
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     8290..8306
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    8762..8778
                     /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 ccgaacttct tgatccttgc ggctacatca gccctgagag
     1021 ccctgtggtt cagctgcaca gcaatttcac cgccgtgtgc gtgctgaaag aaaagtgcat
     1081 ggactacttc cacgtgaacg ccaactacat cgtgtggaaa acgaaccact tcacgatccc
     1141 caaagagcag tacaccatca tcaaccggac cgccagcagc gtgaccttca cagatatcgc
     1201 cagcctgaac atccagctga cctgcaacat cctgaccttc ggccagctgg aacagaacgt
     1261 gtacggcatc acaatcatca gcggcctgcc tccagagaag cccaagaacc tgagctgcat
     1321 cgtgaacgag ggcaagaaaa tgagatgcga gtgggacggc ggcagagaga cacacctgga
     1381 aacaaacttc accctgaagt ccgagtgggc cacacacaag ttcgccgact gcaaggccaa
     1441 gagggatacc cctaccagct gcaccgtgga ctacagcacc gtgtacttcg tgaacatcga
     1501 agtgtgggtc gaagccgaga acgccctggg caaagtgacc agcgaccaca tcaacttcga
     1561 ccccgtgtac aaagtgaagc ccaatcctcc tcacaacctg agcgtgatca acagcgagga
     1621 actgagcagc atcctgaagc tgacatggac aaaccccagc atcaagtccg tgatcatcct
     1681 caagtacaac atccagtacc ggaccaagga cgccagcacc tggtcacaga ttcctccaga
     1741 ggataccgcc tccaccagat ccagcttcac agtgcaggac ctgaagcctt tcaccgagta
     1801 cgtgttccgg atcagatgca tgaaggaaga tggcaaaggc tattggagtg attggagcga
     1861 ggaagccagc ggcatcacct acgaggacag accttctaag gcccctagct tctggtacaa
     1921 gatcgacccc tctcacaccc aaggctacag aaccgtgcag ctcgtctgga aaaccctgcc
     1981 acctttcgag gccaacggca agatcctgga ctacgaagtg accctgacca gatggaagtc
     2041 ccatctgcag aactacaccg tgaatgccac caagctgacc gtgaacctga ccaacgacag
     2101 atacctggcc acactgaccg tgcggaatct cgtgggaaaa tccgatgccg ccgtgctgac
     2161 aatccccgcc tgcgattttc aagccacaca tcccgtgatg gatctgaagg cctttcctaa
     2221 ggacaacatg ctgtgggttg agtggaccac acctcgcgag agcgtgaaaa agtacatcct
     2281 cgagtggtgc gtcctgagcg acaaggcccc ttgtatcacc gactggcagc aagaggatgg
     2341 caccgtgcac agaacctacc tgagaggcaa tctggccgag agcaagtgct acctgatcac
     2401 cgtgacaccc gtgtacgccg atggacctgg aagcccagag agcatcaagg cctacctgaa
     2461 acaggcccct cctagcaagg gccctacagt gcggacaaag aaagtgggca agaacgaggc
     2521 cgtgctggaa tgggatcagc tgcctgtgga tgtgcagaac ggcttcatca gaaactacac
     2581 catcttctac aggaccatca tcggcaacga gacagccgtg aacgtggaca gcagccacac
     2641 agagtacacc ctgagcagcc tgacaagcga caccctgtac atggttcgaa tggccgccta
     2701 caccgacgaa ggcggaaaag atggccccga gttcaccttc accactccta aattcgctca
     2761 gggcgagatc gaaggcgcgc cggaagatca agttgatcca agattgattg atggcaaggg
     2821 cggatctctg gaggtgctgt tccagggacc aggcggaagc ggcaatcccg gcggcgatct
     2881 ggccccccag atgctgcgcg agctgcagga gaccaatgcc gccctgcagg acgtgcgcga
     2941 gctgctgcgc caccgcgtga aggagatcac cttcctgaag aacaccgtga tggagtgcga
     3001 cgcctgcggt atgcagccct ctagaggctc cggctccatc atcccagttg aggaggagaa
     3061 cccggacttc tggaaccgcg aggcagccga ggccctgggt gccgccaaga agctgcagcc
     3121 tgcacagaca gccgccaaga acctcatcat cttcctgggc gatgggatgg gggtgtctac
     3181 ggtgacagct gccaggatcc taaaagggca gaagaaggac aaactggggc ctgagatacc
     3241 cctggccatg gaccgcttcc catatgtggc tctgtccaag acatacaatg tagacaaaca
     3301 tgtgccagac agtggagcca cagccacggc ctacctgtgc ggggtcaagg gcaacttcca
     3361 gaccattggc ttgagtgcag ccgcccgctt taaccagtgc aacacgacac gcggcaacga
     3421 ggtcatctcc gtgatgaatc gggccaagaa agcagggaag tcagtgggag tggtaaccac
     3481 cacacgagtg cagcacgcct cgccagccgg cacctacgcc cacacggtga accgcaactg
     3541 gtactcggac gccgacgtgc ctgcctcggc ccgccaggag gggtgccagg acatcgctac
     3601 gcagctcatc tccaacatgg acattgacgt gatcctaggt ggaggccgaa agtacatgtt
     3661 tcgcatggga accccagacc ctgagtaccc agatgactac agccaaggtg ggaccaggct
     3721 ggacgggaag aatctggtgc aggaatggct ggcgaagcgc cagggtgccc ggtatgtgtg
     3781 gaaccgcact gagctcatgc aggcttccct ggacccgtct gtgacccatc tcatgggtct
     3841 ctttgagcct ggagacatga aatacgagat ccaccgagac tccacactgg acccctccct
     3901 gatggagatg acagaggctg ccctgcgcct gctgagcagg aacccccgcg gcttcttcct
     3961 cttcgtggag ggtggtcgca tcgaccatgg tcatcatgaa agcagggctt accgggcact
     4021 gactgagacg atcatgttcg acgacgccat tgagagggcg ggccagctca ccagcgagga
     4081 ggacacgctg agcctcgtca ctgccgacca ctcccacgtc ttctccttcg gaggctaccc
     4141 cctgcgaggg agctccatct tcgggctggc ccctggcaag gcccgggaca ggaaggccta
     4201 cacggtcctc ctatacggaa acggtccagg ctatgtgctc aaggacggcg cccggccgga
     4261 tgttaccgag agcgagagcg ggagccccga gtatcggcag cagtcagcag tgcccctgga
     4321 cgaagagacc cacgcaggcg aggacgtggc ggtgttcgcg cgcggcccgc aggcgcacct
     4381 ggttcacggc gtgcaggagc agaccttcat agcgcacgtc atggccttcg ccgcctgcct
     4441 ggagccctac accgcctgcg acctggcgcc ccccgccggc accaccgacg ccgcgcaccc
     4501 gggtggcgga ggactgaacg acatcttcga ggctcagaaa atcgaatggc acgaacatca
     4561 ccatcaccat caccatcacc attaaggccg gcctgtagtc actggcacaa cgcgttaccc
     4621 ggtaagccaa tcgggtatac acggtcgtca tactgcagac agggttcttc tactttgcaa
     4681 gatagtcttg agtagtaaaa taaatagata gagaaaatca gcctcgactg tgccttctag
     4741 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     4801 tcccactgtc ctttcctaat aaaatgagga aattgcatca caacactcaa ccctatctcg
     4861 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     4921 ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg
     4981 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     5041 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     5101 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     5161 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     5221 aggccgcctc tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     5281 gcttttgcaa aaagctcccg ggagcttgta tatccatttt cggatctgat cagcacgtgt
     5341 tgacaattaa tcatcggcat agtatatcgg catagtataa tacgacaagg tgaggaacta
     5401 aaccatgacc gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg tccccagggc
     5461 cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca ccgtcgatcc
     5521 ggaccgccac atcgagcggg tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct
     5581 cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg gtggcggtct ggaccacgcc
     5641 ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc ccgcgcatgg ccgagttgag
     5701 cggttcccgg ctggccgcgc agcaacagat ggaaggcctc ctggcgccgc accggcccaa
     5761 ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc gaccaccagg gcaagggtct
     5821 gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag cgcgccgggg tgcccgcctt
     5881 cctggagaca tccgcgcccc gcaacctccc cttctacgag cggctcggct tcaccgtcac
     5941 cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc atgacccgca agcccggtgc
     6001 ctgacacgtg ctacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg
     6061 aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt
     6121 cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     6181 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     6241 catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc
     6301 atggtcatta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     6361 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     6421 ctggccccag cgctgcgatg ataccgcgag aaccacgctc accggctccg gatttatcag
     6481 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     6541 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     6601 tgcgcaacgt tgttgccatc gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     6661 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     6721 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     6781 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     6841 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     6901 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     6961 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     7021 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     7081 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     7141 gggcgacacg gaaatgttga atactcatat tcttcctttt tcaatattat tgaagcattt
     7201 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     7261 taggggtcag tgttacaacc aattaaccaa ttctgaacat tatcgcgagc ccatttatac
     7321 ctgaatatgg ctcataacac cccttgctca tgaccaaaat cccttaacgt gagttacgcg
     7381 cgcgtcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     7441 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     7501 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     7561 gcagatacca aatactgttc ttctagtgta gccgtagtta gcccaccact tcaagaactc
     7621 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     7681 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     7741 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     7801 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     7861 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     7921 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     7981 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     8041 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     8101 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     8161 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa ggcgagagta gggaactgcc
     8221 aggcatcaaa ctaagcagaa ggcccctgac ggatggcctt tttgcgtttc tacaaactct
     8281 ttctgtgttg taaaacgacg gccagtctta agctcgggcc ccctgggcgg ttctgataac
     8341 gagtaatcgt taatccgcaa ataacgtaaa aacccgcttc ggcgggtttt tttatggggg
     8401 gagtttaggg aaagagcatt tgtcagaata tttaagggcg cctgtcactt tgcttgatat
     8461 atgagaatta tttaacctta taaatgagaa aaaagcaacg cactttaaat aagatacgtt
     8521 gctttttcga ttgatgaaca cctataatta aactattcat ctattattta tgattttttg
     8581 tatatacaat atttctagtt tgttaaagag aattaagaaa ataaatctcg aaaataataa
     8641 agggaaaatc agtttttgat atcaaaatta tacatgtcaa cgataataca aaatataata
     8701 caaactataa gatgttatca gtatttatta tcatttagaa taaattttgt gtcgccctta
     8761 attgtgagcg gataacaatt acgagcttca tgcacagtgg cgttgactca atattggcca
     8821 ttagccatat tattcattgg ttatatagca taaatcaata ttggctattg gccattgcat
     8881 acgttgtatc tatatcataa tatgtacatt tatattggct catgtccaat atgaccgcca
     8941 tgtt
//
