LOCUS       sequence-434525-        7645 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 7645)
  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 7645)
  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-TNFRSF14-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..7645
                     /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             3250..3285
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             3295..3318
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             4978..5022
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5023..5049
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(5189..5206)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5195..5306
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5424..5444)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5429..5758
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5609..5744
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5671..5690
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        5806..5853
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5872..6471
                     /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(5872..5891)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6368..6388
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6601..6722
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6638..6657)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6692..6711
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(6775..7566,7567..7635))
                     /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     7398..7417
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(7636..7645,1..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 cggccgcctt gcctagctgc
     2761 aaagaggatg agtatcctgt cggcagcgag tgctgcccta agtgttctcc cggctacaga
     2821 gtgaaagagg cctgcggaga actgaccggc accgtgtgtg aaccttgtcc tcctggcaca
     2881 tatatcgccc acctgaacgg cctgagcaag tgcctgcagt gccagatgtg tgatcctgcc
     2941 atgggactga gagccagcag aaactgcagc agaaccgaga atgccgtgtg tggctgtagc
     3001 cctggccact tctgtatcgt gcaggatggc gatcactgtg ccgcctgtag agcctatgcc
     3061 acatctagcc ctggacagag agtgcagaaa ggcggcaccg agagccagga taccctgtgt
     3121 caaaattgcc ctccaggcac attcagcccc aacggcacac tggaagagtg ccagcaccag
     3181 accaagtgta gctggctggt tacaaaggct ggcgccggaa caagcagcag ccattgggtt
     3241 ggcgcgccgg aagatcaagt tgatccaaga ttgattgatg gcaagggcgg atctctggag
     3301 gtgctgttcc agggaccagg cggaagcggc aatcccggcg gcgatctggc cccccagatg
     3361 ctgcgcgagc tgcaggagac caatgccgcc ctgcaggacg tgcgcgagct gctgcgccac
     3421 cgcgtgaagg agatcacctt cctgaagaac accgtgatgg agtgcgacgc ctgcggtatg
     3481 cagccctcta gaggctccgg ctccatcatc ccagttgagg aggagaaccc ggacttctgg
     3541 aaccgcgagg cagccgaggc cctgggtgcc gccaagaagc tgcagcctgc acagacagcc
     3601 gccaagaacc tcatcatctt cctgggcgat gggatggggg tgtctacggt gacagctgcc
     3661 aggatcctaa aagggcagaa gaaggacaaa ctggggcctg agatacccct ggccatggac
     3721 cgcttcccat atgtggctct gtccaagaca tacaatgtag acaaacatgt gccagacagt
     3781 ggagccacag ccacggccta cctgtgcggg gtcaagggca acttccagac cattggcttg
     3841 agtgcagccg cccgctttaa ccagtgcaac acgacacgcg gcaacgaggt catctccgtg
     3901 atgaatcggg ccaagaaagc agggaagtca gtgggagtgg taaccaccac acgagtgcag
     3961 cacgcctcgc cagccggcac ctacgcccac acggtgaacc gcaactggta ctcggacgcc
     4021 gacgtgcctg cctcggcccg ccaggagggg tgccaggaca tcgctacgca gctcatctcc
     4081 aacatggaca ttgacgtgat cctaggtgga ggccgaaagt acatgtttcg catgggaacc
     4141 ccagaccctg agtacccaga tgactacagc caaggtggga ccaggctgga cgggaagaat
     4201 ctggtgcagg aatggctggc gaagcgccag ggtgcccggt atgtgtggaa ccgcactgag
     4261 ctcatgcagg cttccctgga cccgtctgtg acccatctca tgggtctctt tgagcctgga
     4321 gacatgaaat acgagatcca ccgagactcc acactggacc cctccctgat ggagatgaca
     4381 gaggctgccc tgcgcctgct gagcaggaac ccccgcggct tcttcctctt cgtggagggt
     4441 ggtcgcatcg accatggtca tcatgaaagc agggcttacc gggcactgac tgagacgatc
     4501 atgttcgacg acgccattga gagggcgggc cagctcacca gcgaggagga cacgctgagc
     4561 ctcgtcactg ccgaccactc ccacgtcttc tccttcggag gctaccccct gcgagggagc
     4621 tccatcttcg ggctggcccc tggcaaggcc cgggacagga aggcctacac ggtcctccta
     4681 tacggaaacg gtccaggcta tgtgctcaag gacggcgccc ggccggatgt taccgagagc
     4741 gagagcggga gccccgagta tcggcagcag tcagcagtgc ccctggacga agagacccac
     4801 gcaggcgagg acgtggcggt gttcgcgcgc ggcccgcagg cgcacctggt tcacggcgtg
     4861 caggagcaga ccttcatagc gcacgtcatg gccttcgccg cctgcctgga gccctacacc
     4921 gcctgcgacc tggcgccccc cgccggcacc accgacgccg cgcacccggg tggcggagga
     4981 ctgaacgaca tcttcgaggc tcagaaaatc gaatggcacg aacatcacca tcaccatcac
     5041 catcaccatt aaggccggcc tgtagtcact ggcacaacgc gttacccggt aagccaatcg
     5101 ggtatacacg gtcgtcatac tgcagacagg gttcttctac tttgcaagat agtcttgagt
     5161 agtaaaataa atagatagag aaaatcagcc tcgactgtgc cttctagttg ccagccatct
     5221 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt
     5281 tcctaataaa atgaggaaat tgcatcacaa cactcaaccc tatctcggtc tattcttttg
     5341 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     5401 aatttaacgc gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
     5461 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     5521 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     5581 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     5641 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc
     5701 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     5761 gctcccggga gcttgtatat ccattttcgg atctgatcag cacgtgttga caattaatca
     5821 tcggcatagt atatcggcat agtataatac gacaaggtga ggaactaaac catgaccgag
     5881 tacaagccca cggtgcgcct cgccacccgc gacgacgtcc ccagggccgt acgcaccctc
     5941 gccgccgcgt tcgccgacta ccccgccacg cgccacaccg tcgatccgga ccgccacatc
     6001 gagcgggtca ccgagctgca agaactcttc ctcacgcgcg tcgggctcga catcggcaag
     6061 gtgtgggtcg cggacgacgg cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa
     6121 gcgggggcgg tgttcgccga gatcggcccg cgcatggccg agttgagcgg ttcccggctg
     6181 gccgcgcagc aacagatgga aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg
     6241 ttcctggcca ccgtcggcgt ctcgcccgac caccagggca agggtctggg cagcgccgtc
     6301 gtgctccccg gagtggaggc ggccgagcgc gccggggtgc ccgccttcct ggagacatcc
     6361 gcgccccgca acctcccctt ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag
     6421 gtgcccgaag gaccgcgcac ctggtgcatg acccgcaagc ccggtgcctg acacgtgcta
     6481 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg
     6541 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc
     6601 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     6661 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     6721 tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcattacca
     6781 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     6841 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagcgc
     6901 tgcgatgata ccgcgagaac cacgctcacc ggctccggat ttatcagcaa taaaccagcc
     6961 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     7021 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     7081 tgccatcgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     7141 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     7201 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     7261 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     7321 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     7381 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     7441 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     7501 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     7561 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     7621 atgttgaata ctcatattct tcctt
//
