LOCUS       sequence-303953-        8455 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 8455)
  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 8455)
  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-MIR cl-10-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8455
                     /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             4060..4095
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             4105..4128
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             5788..5832
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5833..5859
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(5999..6016)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6005..6116
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6234..6254)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6239..6568
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6419..6554
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6481..6500
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6616..6663
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6682..7281
                     /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(6682..6701)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7178..7198
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7411..7532
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7448..7467)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7502..7521
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7585..8376,8377..8445))
                     /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     8208..8227
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8446..8455,1..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 cggccgccgg aacaattcct
     2761 aagcctacac tgtgggccga gcctgacagc gtgatcacac agggatctcc cgtgacactg
     2821 agctgtcagg gaagcctgga agcccaagag taccggctgt acagagagaa gaagtccgcc
     2881 agctggatca cccggatcag acctgagctg gtcaagaacg gccagtttca catccccagc
     2941 atcacctggg agcacaccgg cagatacggc tgccagtact atagcagagc ccgttggagc
     3001 gagctgagcg atcctctggt gctggttatg accggcgctt accccaagcc aacactgtct
     3061 gctcagccct ctccagtggt cacatctggc ggaagagtga ccctgcagtg cgaaagccag
     3121 gtggcctttg gcggcttcat cctgtgcaaa gaaggcgagg acgagcaccc tcagtgcctg
     3181 aattctcagc ctcacgccag aggaagcagc cgggccattt tttctgtggg ccccgtgtct
     3241 cccaacagac ggtggtccca cagatgctac ggctacgacc tgaacagccc ctacgtgtgg
     3301 tctagcccta gcgacctgct ggaactgctg gttcctggcg tgtccaagaa acccagcctg
     3361 tctgtgcagc ccggacctgt tatggctcct ggcgaatctc tgactctgca gtgtgtgtcc
     3421 gacgtgggct acgacagatt cgtgctctac aaagagggcg agcgcgacct cagacaactg
     3481 cctggtagac agcctcaggc cggactgtct caggccaatt tcacactggg ccctgtgtcc
     3541 agaagctatg gcggccagta cagatgttat ggcgcccaca atctgagcag cgagtgtagc
     3601 gctcctagcg atcccctgga catcctgatc accggccaga tcagaggcac ccctttcatc
     3661 agtgtccagc ctggacctac agtggccagc ggcgaaaatg tgacactgct gtgccagagc
     3721 tggcggcagt tccacacctt tctgctgaca aaagccggcg ctgccgacgc tcctctgaga
     3781 ctgagatcta tccacgagta ccccaagtac caggccgagt ttcctatgag ccctgtgaca
     3841 tctgcccacg ccggcaccta tagatgttac ggcagcctga atagcgaccc ctacctgctg
     3901 tctcacccta gcgaaccact cgagctggtg gtgtctggcc cttctatggg aagctcccca
     3961 cctccaacag gccctatctc tacacctgga cctgaggacc agcctctgac accaacaggc
     4021 tctgatcctc agtctggcct gggcagacat ggcgcgccgg aagatcaagt tgatccaaga
     4081 ttgattgatg gcaagggcgg atctctggag gtgctgttcc agggaccagg cggaagcggc
     4141 aatcccggcg gcgatctggc cccccagatg ctgcgcgagc tgcaggagac caatgccgcc
     4201 ctgcaggacg tgcgcgagct gctgcgccac cgcgtgaagg agatcacctt cctgaagaac
     4261 accgtgatgg agtgcgacgc ctgcggtatg cagccctcta gaggctccgg ctccatcatc
     4321 ccagttgagg aggagaaccc ggacttctgg aaccgcgagg cagccgaggc cctgggtgcc
     4381 gccaagaagc tgcagcctgc acagacagcc gccaagaacc tcatcatctt cctgggcgat
     4441 gggatggggg tgtctacggt gacagctgcc aggatcctaa aagggcagaa gaaggacaaa
     4501 ctggggcctg agatacccct ggccatggac cgcttcccat atgtggctct gtccaagaca
     4561 tacaatgtag acaaacatgt gccagacagt ggagccacag ccacggccta cctgtgcggg
     4621 gtcaagggca acttccagac cattggcttg agtgcagccg cccgctttaa ccagtgcaac
     4681 acgacacgcg gcaacgaggt catctccgtg atgaatcggg ccaagaaagc agggaagtca
     4741 gtgggagtgg taaccaccac acgagtgcag cacgcctcgc cagccggcac ctacgcccac
     4801 acggtgaacc gcaactggta ctcggacgcc gacgtgcctg cctcggcccg ccaggagggg
     4861 tgccaggaca tcgctacgca gctcatctcc aacatggaca ttgacgtgat cctaggtgga
     4921 ggccgaaagt acatgtttcg catgggaacc ccagaccctg agtacccaga tgactacagc
     4981 caaggtggga ccaggctgga cgggaagaat ctggtgcagg aatggctggc gaagcgccag
     5041 ggtgcccggt atgtgtggaa ccgcactgag ctcatgcagg cttccctgga cccgtctgtg
     5101 acccatctca tgggtctctt tgagcctgga gacatgaaat acgagatcca ccgagactcc
     5161 acactggacc cctccctgat ggagatgaca gaggctgccc tgcgcctgct gagcaggaac
     5221 ccccgcggct tcttcctctt cgtggagggt ggtcgcatcg accatggtca tcatgaaagc
     5281 agggcttacc gggcactgac tgagacgatc atgttcgacg acgccattga gagggcgggc
     5341 cagctcacca gcgaggagga cacgctgagc ctcgtcactg ccgaccactc ccacgtcttc
     5401 tccttcggag gctaccccct gcgagggagc tccatcttcg ggctggcccc tggcaaggcc
     5461 cgggacagga aggcctacac ggtcctccta tacggaaacg gtccaggcta tgtgctcaag
     5521 gacggcgccc ggccggatgt taccgagagc gagagcggga gccccgagta tcggcagcag
     5581 tcagcagtgc ccctggacga agagacccac gcaggcgagg acgtggcggt gttcgcgcgc
     5641 ggcccgcagg cgcacctggt tcacggcgtg caggagcaga ccttcatagc gcacgtcatg
     5701 gccttcgccg cctgcctgga gccctacacc gcctgcgacc tggcgccccc cgccggcacc
     5761 accgacgccg cgcacccggg tggcggagga ctgaacgaca tcttcgaggc tcagaaaatc
     5821 gaatggcacg aacatcacca tcaccatcac catcaccatt aaggccggcc tgtagtcact
     5881 ggcacaacgc gttacccggt aagccaatcg ggtatacacg gtcgtcatac tgcagacagg
     5941 gttcttctac tttgcaagat agtcttgagt agtaaaataa atagatagag aaaatcagcc
     6001 tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg
     6061 accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcacaa
     6121 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     6181 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt
     6241 gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     6301 gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag
     6361 tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat
     6421 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt
     6481 tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg
     6541 cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg
     6601 atctgatcag cacgtgttga caattaatca tcggcatagt atatcggcat agtataatac
     6661 gacaaggtga ggaactaaac catgaccgag tacaagccca cggtgcgcct cgccacccgc
     6721 gacgacgtcc ccagggccgt acgcaccctc gccgccgcgt tcgccgacta ccccgccacg
     6781 cgccacaccg tcgatccgga ccgccacatc gagcgggtca ccgagctgca agaactcttc
     6841 ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg cggacgacgg cgccgcggtg
     6901 gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga gatcggcccg
     6961 cgcatggccg agttgagcgg ttcccggctg gccgcgcagc aacagatgga aggcctcctg
     7021 gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca ccgtcggcgt ctcgcccgac
     7081 caccagggca agggtctggg cagcgccgtc gtgctccccg gagtggaggc ggccgagcgc
     7141 gccggggtgc ccgccttcct ggagacatcc gcgccccgca acctcccctt ctacgagcgg
     7201 ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac ctggtgcatg
     7261 acccgcaagc ccggtgcctg acacgtgcta cgagatttcg attccaccgc cgccttctat
     7321 gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg
     7381 gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac
     7441 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
     7501 tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc
     7561 tagagcttgg cgtaatcatg gtcattacca atgcttaatc agtgaggcac ctatctcagc
     7621 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     7681 acgggagggc ttaccatctg gccccagcgc tgcgatgata ccgcgagaac cacgctcacc
     7741 ggctccggat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     7801 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     7861 ttcgccagtt aatagtttgc gcaacgttgt tgccatcgct acaggcatcg tggtgtcacg
     7921 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     7981 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     8041 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     8101 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     8161 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     8221 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     8281 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     8341 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     8401 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatattct tcctt
//
