LOCUS       sequence-400999-        8191 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 8191)
  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 8191)
  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-IL6R-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8191
                     /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             2030..2065
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             2075..2098
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             3758..3802
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             3803..3829
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(3969..3986)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3975..4086
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4204..4224)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4209..4538
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4389..4524
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4451..4470
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4586..4633
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4652..5251
                     /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(4652..4671)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5148..5168
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5381..5502
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5418..5437)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5472..5491
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(5555..6346,6347..6415))
                     /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     6178..6197
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6416..6507)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6549..6568
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6574..6593
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      6679..7267
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7168..7187
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7421..7438
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     7536..7553
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7537..7553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    8009..8025
                     /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 ccttggctcc tagaagatgt cctgctcaag aggtggccag
     1021 aggcgtgttg acatctctgc ctggcgatag cgtgaccctg acatgtcctg gcgtggaacc
     1081 tgaggataat gccaccgtgc actgggtgct gagaaaacct gctgccggat ctcatcctag
     1141 cagatgggcc ggaatgggcc gtagactgct gctgagatct gtccagctgc acgacagcgg
     1201 caactacagc tgttatagag ccggcagacc agccggcaca gtgcatctgc ttgttgacgt
     1261 gccacctgag gaaccccagc tgagctgctt tagaaagagc cctctgagca acgtcgtgtg
     1321 cgagtgggga cctagaagca cccctagcct gacaacaaaa gccgtgctgc tcgtgcggaa
     1381 gttccagaat agccctgccg aggacttcca agagccttgc cagtacagcc aagagagcca
     1441 gaagttcagc tgtcagctgg ccgtgcctga gggcgatagc agcttctaca tcgtgtctat
     1501 gtgcgtggcc agcagcgtgg gcagcaagtt tagcaagacc cagaccttcc aaggctgcgg
     1561 cattctgcaa cccgatcctc cagccaatat caccgtgacc gccgtggcca gaaatcccag
     1621 atggctgtct gtgacctggc aggaccctca cagctggaac agcagctttt accggctgag
     1681 attcgagctg cggtacagag ccgagagaag caagaccttc accacctgga tggtcaagga
     1741 cctgcagcac cactgcgtga tccatgatgc ttggagcgga ctgagacacg tggtgcagct
     1801 gagagcccaa gaggaatttg gacaaggcga gtggtccgaa tggtcccctg aagctatggg
     1861 caccccttgg acagagagca gaagccctcc tgccgagaat gaggtgtcaa cacctatgca
     1921 ggccctgacc accaacaagg acgacgacaa catcctgttc agagacagcg ccaatgccac
     1981 cagcctgcct gtgcaggatt ctagctctgt gcctctgcct ggcgcgccgg aagatcaagt
     2041 tgatccaaga ttgattgatg gcaagggcgg atctctggag gtgctgttcc agggaccagg
     2101 cggaagcggc aatcccggcg gcgatctggc cccccagatg ctgcgcgagc tgcaggagac
     2161 caatgccgcc ctgcaggacg tgcgcgagct gctgcgccac cgcgtgaagg agatcacctt
     2221 cctgaagaac accgtgatgg agtgcgacgc ctgcggtatg cagccctcta gaggctccgg
     2281 ctccatcatc ccagttgagg aggagaaccc ggacttctgg aaccgcgagg cagccgaggc
     2341 cctgggtgcc gccaagaagc tgcagcctgc acagacagcc gccaagaacc tcatcatctt
     2401 cctgggcgat gggatggggg tgtctacggt gacagctgcc aggatcctaa aagggcagaa
     2461 gaaggacaaa ctggggcctg agatacccct ggccatggac cgcttcccat atgtggctct
     2521 gtccaagaca tacaatgtag acaaacatgt gccagacagt ggagccacag ccacggccta
     2581 cctgtgcggg gtcaagggca acttccagac cattggcttg agtgcagccg cccgctttaa
     2641 ccagtgcaac acgacacgcg gcaacgaggt catctccgtg atgaatcggg ccaagaaagc
     2701 agggaagtca gtgggagtgg taaccaccac acgagtgcag cacgcctcgc cagccggcac
     2761 ctacgcccac acggtgaacc gcaactggta ctcggacgcc gacgtgcctg cctcggcccg
     2821 ccaggagggg tgccaggaca tcgctacgca gctcatctcc aacatggaca ttgacgtgat
     2881 cctaggtgga ggccgaaagt acatgtttcg catgggaacc ccagaccctg agtacccaga
     2941 tgactacagc caaggtggga ccaggctgga cgggaagaat ctggtgcagg aatggctggc
     3001 gaagcgccag ggtgcccggt atgtgtggaa ccgcactgag ctcatgcagg cttccctgga
     3061 cccgtctgtg acccatctca tgggtctctt tgagcctgga gacatgaaat acgagatcca
     3121 ccgagactcc acactggacc cctccctgat ggagatgaca gaggctgccc tgcgcctgct
     3181 gagcaggaac ccccgcggct tcttcctctt cgtggagggt ggtcgcatcg accatggtca
     3241 tcatgaaagc agggcttacc gggcactgac tgagacgatc atgttcgacg acgccattga
     3301 gagggcgggc cagctcacca gcgaggagga cacgctgagc ctcgtcactg ccgaccactc
     3361 ccacgtcttc tccttcggag gctaccccct gcgagggagc tccatcttcg ggctggcccc
     3421 tggcaaggcc cgggacagga aggcctacac ggtcctccta tacggaaacg gtccaggcta
     3481 tgtgctcaag gacggcgccc ggccggatgt taccgagagc gagagcggga gccccgagta
     3541 tcggcagcag tcagcagtgc ccctggacga agagacccac gcaggcgagg acgtggcggt
     3601 gttcgcgcgc ggcccgcagg cgcacctggt tcacggcgtg caggagcaga ccttcatagc
     3661 gcacgtcatg gccttcgccg cctgcctgga gccctacacc gcctgcgacc tggcgccccc
     3721 cgccggcacc accgacgccg cgcacccggg tggcggagga ctgaacgaca tcttcgaggc
     3781 tcagaaaatc gaatggcacg aacatcacca tcaccatcac catcaccatt aaggccggcc
     3841 tgtagtcact ggcacaacgc gttacccggt aagccaatcg ggtatacacg gtcgtcatac
     3901 tgcagacagg gttcttctac tttgcaagat agtcttgagt agtaaaataa atagatagag
     3961 aaaatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt
     4021 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat
     4081 tgcatcacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     4141 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc
     4201 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     4261 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     4321 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     4381 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     4441 taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt
     4501 agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat
     4561 ccattttcgg atctgatcag cacgtgttga caattaatca tcggcatagt atatcggcat
     4621 agtataatac gacaaggtga ggaactaaac catgaccgag tacaagccca cggtgcgcct
     4681 cgccacccgc gacgacgtcc ccagggccgt acgcaccctc gccgccgcgt tcgccgacta
     4741 ccccgccacg cgccacaccg tcgatccgga ccgccacatc gagcgggtca ccgagctgca
     4801 agaactcttc ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg cggacgacgg
     4861 cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga
     4921 gatcggcccg cgcatggccg agttgagcgg ttcccggctg gccgcgcagc aacagatgga
     4981 aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca ccgtcggcgt
     5041 ctcgcccgac caccagggca agggtctggg cagcgccgtc gtgctccccg gagtggaggc
     5101 ggccgagcgc gccggggtgc ccgccttcct ggagacatcc gcgccccgca acctcccctt
     5161 ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac
     5221 ctggtgcatg acccgcaagc ccggtgcctg acacgtgcta cgagatttcg attccaccgc
     5281 cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct
     5341 ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta
     5401 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     5461 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc
     5521 gacctctagc tagagcttgg cgtaatcatg gtcattacca atgcttaatc agtgaggcac
     5581 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     5641 taactacgat acgggagggc ttaccatctg gccccagcgc tgcgatgata ccgcgagaac
     5701 cacgctcacc ggctccggat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     5761 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     5821 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccatcgct acaggcatcg
     5881 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     5941 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     6001 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     6061 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     6121 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     6181 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     6241 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     6301 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     6361 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatattct
     6421 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     6481 ttgaatgtat ttagaaaaat aaacaaatag gggtcagtgt tacaaccaat taaccaattc
     6541 tgaacattat cgcgagccca tttatacctg aatatggctc ataacacccc ttgctcatga
     6601 ccaaaatccc ttaacgtgag ttacgcgcgc gtcgttccac tgagcgtcag accccgtaga
     6661 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     6721 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     6781 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     6841 gtagttagcc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     6901 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     6961 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     7021 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     7081 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     7141 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     7201 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     7261 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     7321 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
     7381 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
     7441 agcggaaggc gagagtaggg aactgccagg catcaaacta agcagaaggc ccctgacgga
     7501 tggccttttt gcgtttctac aaactctttc tgtgttgtaa aacgacggcc agtcttaagc
     7561 tcgggccccc tgggcggttc tgataacgag taatcgttaa tccgcaaata acgtaaaaac
     7621 ccgcttcggc gggttttttt atggggggag tttagggaaa gagcatttgt cagaatattt
     7681 aagggcgcct gtcactttgc ttgatatatg agaattattt aaccttataa atgagaaaaa
     7741 agcaacgcac tttaaataag atacgttgct ttttcgattg atgaacacct ataattaaac
     7801 tattcatcta ttatttatga ttttttgtat atacaatatt tctagtttgt taaagagaat
     7861 taagaaaata aatctcgaaa ataataaagg gaaaatcagt ttttgatatc aaaattatac
     7921 atgtcaacga taatacaaaa tataatacaa actataagat gttatcagta tttattatca
     7981 tttagaataa attttgtgtc gcccttaatt gtgagcggat aacaattacg agcttcatgc
     8041 acagtggcgt tgactcaata ttggccatta gccatattat tcattggtta tatagcataa
     8101 atcaatattg gctattggcc attgcatacg ttgtatctat atcataatat gtacatttat
     8161 attggctcat gtccaatatg accgccatgt t
//
