LOCUS       sequence-304015-        8662 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 8662)
  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 8662)
  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-DLL4-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8662
                     /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             4267..4302
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             4312..4335
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             5995..6039
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6040..6066
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(6206..6223)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6212..6323
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6441..6461)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6446..6775
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6626..6761
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6688..6707
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6823..6870
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6889..7488
                     /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(6889..6908)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7385..7405
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7618..7739
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7655..7674)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7709..7728
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7792..8583,8584..8652))
                     /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     8415..8434
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8653..8662,1..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 cggagttttt
     2761 cagctgcaac tgcaagagtt catcaacgag agaggcgtgc tggcctctgg cagaccttgt
     2821 gaacctggct gccggacctt cttcagagtg tgcctgaagc acttccaggc cgtggtgtct
     2881 cctggacctt gtacctttgg caccgtgtct acacctgtgc tgggcaccaa tagcttcgcc
     2941 gtgcgggatg attctagcgg cggaggcaga aatcctctgc agctgccctt caacttcacc
     3001 tggcctggca ccttcagcct gatcatcgaa gcctggcatg cccctgggga tgatctgagg
     3061 cctgaagctt tgcctcctga cgctctgatc agcaagatcg ccatccaggg atctctggcc
     3121 gtgggccaaa attggctgct ggacgagcag accagcacac tgaccagact gcggtacagc
     3181 tacagagtga tctgcagcga caactactac ggcgacaact gcagcagact gtgcaagaag
     3241 cggaacgacc acttcggcca ctacgtgtgt cagcccgatg gcaacctgtc ttgtctgcct
     3301 ggatggaccg gcgagtactg ccagcaacct atctgtctga gcggctgcca cgagcagaat
     3361 ggctactgtt ctaagcccgc cgagtgcctg tgtagacctg gctggcaagg caggctgtgc
     3421 aacgagtgca tccctcacaa tggctgcaga cacggcacct gtagcacccc ttggcagtgc
     3481 acatgtgacg aaggatgggg cggactgttc tgcgaccaag acctgaacta ctgcacccat
     3541 cacagcccct gcaagaacgg cgccacatgt agcaacagcg gccagcggtc ttacacctgt
     3601 acctgcagac ctggatacac cggcgtggac tgcgagctgg aactgagcga gtgcgacagc
     3661 aacccctgta gaaatggcgg ctcctgcaag gaccaagagg acggctatca ctgcctgtgt
     3721 ccacctggat attacggcct gcactgcgaa cacagcaccc tgagctgtgc cgactctccc
     3781 tgttttaatg gcggcagctg tagagagaga aaccagggcg ccaattacgc ctgcgagtgc
     3841 ccacctaatt tcaccggcag caactgcgag aagaaagtgg accggtgcac cagcaatccc
     3901 tgcgctaatg gtggccagtg cctgaacaga ggccccagca gaatgtgtag atgcagaccc
     3961 ggctttaccg gcacctattg cgaactgcac gtgtccgact gcgcccggaa tccttgtgct
     4021 catggcggca catgccacga cctggaaaac ggcctgatgt gcacttgccc tgccggcttc
     4081 tctgggagaa gatgtgaagt gcggaccagc atcgacgcct gtgccagctc tccttgcttc
     4141 aacagagcca cctgttacac cgacctgagc accgacacct tcgtgtgcaa ctgcccttac
     4201 ggcttcgtgg gcagcagatg cgagtttcct gttggcctgc ctcctagctt cccttggggc
     4261 gcgccggaag atcaagttga tccaagattg attgatggca agggcggatc tctggaggtg
     4321 ctgttccagg gaccaggcgg aagcggcaat cccggcggcg atctggcccc ccagatgctg
     4381 cgcgagctgc aggagaccaa tgccgccctg caggacgtgc gcgagctgct gcgccaccgc
     4441 gtgaaggaga tcaccttcct gaagaacacc gtgatggagt gcgacgcctg cggtatgcag
     4501 ccctctagag gctccggctc catcatccca gttgaggagg agaacccgga cttctggaac
     4561 cgcgaggcag ccgaggccct gggtgccgcc aagaagctgc agcctgcaca gacagccgcc
     4621 aagaacctca tcatcttcct gggcgatggg atgggggtgt ctacggtgac agctgccagg
     4681 atcctaaaag ggcagaagaa ggacaaactg gggcctgaga tacccctggc catggaccgc
     4741 ttcccatatg tggctctgtc caagacatac aatgtagaca aacatgtgcc agacagtgga
     4801 gccacagcca cggcctacct gtgcggggtc aagggcaact tccagaccat tggcttgagt
     4861 gcagccgccc gctttaacca gtgcaacacg acacgcggca acgaggtcat ctccgtgatg
     4921 aatcgggcca agaaagcagg gaagtcagtg ggagtggtaa ccaccacacg agtgcagcac
     4981 gcctcgccag ccggcaccta cgcccacacg gtgaaccgca actggtactc ggacgccgac
     5041 gtgcctgcct cggcccgcca ggaggggtgc caggacatcg ctacgcagct catctccaac
     5101 atggacattg acgtgatcct aggtggaggc cgaaagtaca tgtttcgcat gggaacccca
     5161 gaccctgagt acccagatga ctacagccaa ggtgggacca ggctggacgg gaagaatctg
     5221 gtgcaggaat ggctggcgaa gcgccagggt gcccggtatg tgtggaaccg cactgagctc
     5281 atgcaggctt ccctggaccc gtctgtgacc catctcatgg gtctctttga gcctggagac
     5341 atgaaatacg agatccaccg agactccaca ctggacccct ccctgatgga gatgacagag
     5401 gctgccctgc gcctgctgag caggaacccc cgcggcttct tcctcttcgt ggagggtggt
     5461 cgcatcgacc atggtcatca tgaaagcagg gcttaccggg cactgactga gacgatcatg
     5521 ttcgacgacg ccattgagag ggcgggccag ctcaccagcg aggaggacac gctgagcctc
     5581 gtcactgccg accactccca cgtcttctcc ttcggaggct accccctgcg agggagctcc
     5641 atcttcgggc tggcccctgg caaggcccgg gacaggaagg cctacacggt cctcctatac
     5701 ggaaacggtc caggctatgt gctcaaggac ggcgcccggc cggatgttac cgagagcgag
     5761 agcgggagcc ccgagtatcg gcagcagtca gcagtgcccc tggacgaaga gacccacgca
     5821 ggcgaggacg tggcggtgtt cgcgcgcggc ccgcaggcgc acctggttca cggcgtgcag
     5881 gagcagacct tcatagcgca cgtcatggcc ttcgccgcct gcctggagcc ctacaccgcc
     5941 tgcgacctgg cgccccccgc cggcaccacc gacgccgcgc acccgggtgg cggaggactg
     6001 aacgacatct tcgaggctca gaaaatcgaa tggcacgaac atcaccatca ccatcaccat
     6061 caccattaag gccggcctgt agtcactggc acaacgcgtt acccggtaag ccaatcgggt
     6121 atacacggtc gtcatactgc agacagggtt cttctacttt gcaagatagt cttgagtagt
     6181 aaaataaata gatagagaaa atcagcctcg actgtgcctt ctagttgcca gccatctgtt
     6241 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     6301 taataaaatg aggaaattgc atcacaacac tcaaccctat ctcggtctat tcttttgatt
     6361 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     6421 ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc
     6481 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa
     6541 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     6601 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
     6661 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctctgcctc
     6721 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct
     6781 cccgggagct tgtatatcca ttttcggatc tgatcagcac gtgttgacaa ttaatcatcg
     6841 gcatagtata tcggcatagt ataatacgac aaggtgagga actaaaccat gaccgagtac
     6901 aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg caccctcgcc
     6961 gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg ccacatcgag
     7021 cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg
     7081 tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag cgtcgaagcg
     7141 ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc ccggctggcc
     7201 gcgcagcaac agatggaagg cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc
     7261 ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg
     7321 ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga gacatccgcg
     7381 ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg
     7441 cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctgaca cgtgctacga
     7501 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac
     7561 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac
     7621 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     7681 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     7741 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc attaccaatg
     7801 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     7861 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagcgctgc
     7921 gatgataccg cgagaaccac gctcaccggc tccggattta tcagcaataa accagccagc
     7981 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     8041 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     8101 catcgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     8161 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     8221 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     8281 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     8341 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     8401 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     8461 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     8521 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     8581 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     8641 ttgaatactc atattcttcc tt
//
