LOCUS       sequence-304085-        8968 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 8968)
  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 8968)
  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-LRRN1-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8968
                     /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             4573..4608
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             4618..4641
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             6301..6345
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6346..6372
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(6512..6529)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6518..6629
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6747..6767)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6752..7081
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6932..7067
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6994..7013
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7129..7176
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7195..7794
                     /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(7195..7214)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7691..7711
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7924..8045
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7961..7980)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8015..8034
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(8098..8889,8890..8958))
                     /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     8721..8740
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8959..8968,1..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 cggccgccat tcagaatagc
     2761 gagtgccctc agctgtgcgt gtgcgagatc agaccctggt ttacccctca gagcacctac
     2821 agagaagcca ccaccgtgga ctgcaacgac ctgcggctga caagaatccc cagcaacctg
     2881 agcagcgaca cccaggttct gctgctgcag agcaacaata tcgccaagac cgtggacgaa
     2941 ctccagcagc tgttcaacct gaccgagctg gacttcagcc agaacaactt caccaacatc
     3001 aaagaagtcg gcctggccaa tctgacccag ctgacaacac tgcacctgga agagaaccag
     3061 atcaccgaga tgaccgacta ctgcctgcaa gacctgagca acctgcaaga gctgtacatc
     3121 aaccacaacc agatctccac catcagcgcc cacgcctttg ccggcctgaa gaatctgctg
     3181 aggctgcacc tgaacagcaa caagctgaaa gtgatcgaca gccgttggtt cgacagcacc
     3241 cctaacctgg aaatcctgat gatcggcgag aaccccgtga tcggcatcct ggacatgaac
     3301 ttcaagcccc tggctaacct gcggagcctg gtgctggctg gcatgtacct gacagacatc
     3361 cctggcaatg ccctcgtggg cctcgatagc ctggaaagcc tgtccttcta cgacaacaag
     3421 ctcgtgaagg tgccccagct ggccctgcag aaagtgccca atctgaagtt cctggacctc
     3481 aacaagaacc ccatccacaa gatccaagag ggcgacttca agaacatgct gcgcctgaaa
     3541 gagctgggga tcaacaacat gggcgagctg gtgtccgtgg acagatacgc cctggacaat
     3601 ctgcctgagc tgaccaaact ggaagccaca aacaacccca agctgagcta catccaccgg
     3661 ctggccttta gaagcgtgcc cgctctggaa tccctgatgc tgaacaacaa cgccctgaac
     3721 gccatctacc agaaaaccgt ggaatctctg cccaacctga gagagatcag catccacagc
     3781 aaccctctga gatgcgactg cgtgatccac tggattaact ccaacaagac caacatccgg
     3841 ttcatggaac ccctgtccat gttctgcgcc atgcctccag agtacaaggg ccaccaagtg
     3901 aaagaggtgc tgatccagga cagcagcgag cagtgcctgc ctatgatcag ccacgacagc
     3961 ttccccaacc ggctgaatgt ggatatcggc accacagtgt ttctggactg ccgcgccatg
     4021 gccgagcctg agcctgaaat ctattgggtc acccctatcg gcaacaagat caccgtggaa
     4081 accctgagcg acaagtacaa gctgtccagc gagggcacac tggaaatcag caacatccag
     4141 atcgaggaca gcggccggta tacctgcgtg gcacagaatg tgcagggcgc cgatacaaga
     4201 gtggccacca tcaaagtgaa cggcaccctg ctggatggca cccaggtgct gaagatctac
     4261 gtgaagcaga ccgagagcca cagcatcctg gtgtcttgga aagtgaactc caacgtgatg
     4321 acctccaacc tgaagtggtc cagcgccacc atgaagatcg acaaccctca catcacctac
     4381 accgccagag tgcccgtgga tgtgcacgag tacaacctga cacatctgca gcccagcacc
     4441 gactacgaag tgtgcctgac cgtgtccaac atccaccagc agacccagaa aagctgcgtg
     4501 aacgtgacca ccaagaacgc cgccttcgcc gtggacatca gcgaccaaga gacaagcaca
     4561 gctggcgcgc cggaagatca agttgatcca agattgattg atggcaaggg cggatctctg
     4621 gaggtgctgt tccagggacc aggcggaagc ggcaatcccg gcggcgatct ggccccccag
     4681 atgctgcgcg agctgcagga gaccaatgcc gccctgcagg acgtgcgcga gctgctgcgc
     4741 caccgcgtga aggagatcac cttcctgaag aacaccgtga tggagtgcga cgcctgcggt
     4801 atgcagccct ctagaggctc cggctccatc atcccagttg aggaggagaa cccggacttc
     4861 tggaaccgcg aggcagccga ggccctgggt gccgccaaga agctgcagcc tgcacagaca
     4921 gccgccaaga acctcatcat cttcctgggc gatgggatgg gggtgtctac ggtgacagct
     4981 gccaggatcc taaaagggca gaagaaggac aaactggggc ctgagatacc cctggccatg
     5041 gaccgcttcc catatgtggc tctgtccaag acatacaatg tagacaaaca tgtgccagac
     5101 agtggagcca cagccacggc ctacctgtgc ggggtcaagg gcaacttcca gaccattggc
     5161 ttgagtgcag ccgcccgctt taaccagtgc aacacgacac gcggcaacga ggtcatctcc
     5221 gtgatgaatc gggccaagaa agcagggaag tcagtgggag tggtaaccac cacacgagtg
     5281 cagcacgcct cgccagccgg cacctacgcc cacacggtga accgcaactg gtactcggac
     5341 gccgacgtgc ctgcctcggc ccgccaggag gggtgccagg acatcgctac gcagctcatc
     5401 tccaacatgg acattgacgt gatcctaggt ggaggccgaa agtacatgtt tcgcatggga
     5461 accccagacc ctgagtaccc agatgactac agccaaggtg ggaccaggct ggacgggaag
     5521 aatctggtgc aggaatggct ggcgaagcgc cagggtgccc ggtatgtgtg gaaccgcact
     5581 gagctcatgc aggcttccct ggacccgtct gtgacccatc tcatgggtct ctttgagcct
     5641 ggagacatga aatacgagat ccaccgagac tccacactgg acccctccct gatggagatg
     5701 acagaggctg ccctgcgcct gctgagcagg aacccccgcg gcttcttcct cttcgtggag
     5761 ggtggtcgca tcgaccatgg tcatcatgaa agcagggctt accgggcact gactgagacg
     5821 atcatgttcg acgacgccat tgagagggcg ggccagctca ccagcgagga ggacacgctg
     5881 agcctcgtca ctgccgacca ctcccacgtc ttctccttcg gaggctaccc cctgcgaggg
     5941 agctccatct tcgggctggc ccctggcaag gcccgggaca ggaaggccta cacggtcctc
     6001 ctatacggaa acggtccagg ctatgtgctc aaggacggcg cccggccgga tgttaccgag
     6061 agcgagagcg ggagccccga gtatcggcag cagtcagcag tgcccctgga cgaagagacc
     6121 cacgcaggcg aggacgtggc ggtgttcgcg cgcggcccgc aggcgcacct ggttcacggc
     6181 gtgcaggagc agaccttcat agcgcacgtc atggccttcg ccgcctgcct ggagccctac
     6241 accgcctgcg acctggcgcc ccccgccggc accaccgacg ccgcgcaccc gggtggcgga
     6301 ggactgaacg acatcttcga ggctcagaaa atcgaatggc acgaacatca ccatcaccat
     6361 caccatcacc attaaggccg gcctgtagtc actggcacaa cgcgttaccc ggtaagccaa
     6421 tcgggtatac acggtcgtca tactgcagac agggttcttc tactttgcaa gatagtcttg
     6481 agtagtaaaa taaatagata gagaaaatca gcctcgactg tgccttctag ttgccagcca
     6541 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
     6601 ctttcctaat aaaatgagga aattgcatca caacactcaa ccctatctcg gtctattctt
     6661 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     6721 aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     6781 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     6841 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     6901 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     6961 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     7021 tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     7081 aaagctcccg ggagcttgta tatccatttt cggatctgat cagcacgtgt tgacaattaa
     7141 tcatcggcat agtatatcgg catagtataa tacgacaagg tgaggaacta aaccatgacc
     7201 gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg tccccagggc cgtacgcacc
     7261 ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca ccgtcgatcc ggaccgccac
     7321 atcgagcggg tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct cgacatcggc
     7381 aaggtgtggg tcgcggacga cggcgccgcg gtggcggtct ggaccacgcc ggagagcgtc
     7441 gaagcggggg cggtgttcgc cgagatcggc ccgcgcatgg ccgagttgag cggttcccgg
     7501 ctggccgcgc agcaacagat ggaaggcctc ctggcgccgc accggcccaa ggagcccgcg
     7561 tggttcctgg ccaccgtcgg cgtctcgccc gaccaccagg gcaagggtct gggcagcgcc
     7621 gtcgtgctcc ccggagtgga ggcggccgag cgcgccgggg tgcccgcctt cctggagaca
     7681 tccgcgcccc gcaacctccc cttctacgag cggctcggct tcaccgtcac cgccgacgtc
     7741 gaggtgcccg aaggaccgcg cacctggtgc atgacccgca agcccggtgc ctgacacgtg
     7801 ctacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc
     7861 cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac
     7921 cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     7981 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     8041 tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatta
     8101 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     8161 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     8221 cgctgcgatg ataccgcgag aaccacgctc accggctccg gatttatcag caataaacca
     8281 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     8341 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     8401 tgttgccatc gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     8461 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     8521 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     8581 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     8641 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     8701 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     8761 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     8821 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     8881 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     8941 gaaatgttga atactcatat tcttcctt
//
