LOCUS       sequence-400994-        8257 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 8257)
  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 8257)
  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-FCRL4-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8257
                     /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             2096..2131
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             2141..2164
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             3824..3868
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             3869..3895
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(4035..4052)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4041..4152
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4270..4290)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4275..4604
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4455..4590
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4517..4536
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4652..4699
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4718..5317
                     /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(4718..4737)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5214..5234
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5447..5568
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5484..5503)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5538..5557
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(5621..6412,6413..6481))
                     /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     6244..6263
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6482..6573)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6615..6634
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6640..6659
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      6745..7333
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7234..7253
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7487..7504
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     7602..7619
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7603..7619
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    8075..8091
                     /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 ccgctcacaa acctgtgatt tctgtgcacc ctccttggac
     1021 caccttcttc aagggcgaga gagtgaccct gacctgcaac ggcttccagt tctacgccac
     1081 cgagaaaacc acctggtatc accggcacta ctggggcgag aaactgacac tgacacccgg
     1141 caacaccctg gaagtccgcg agtctggact gtacagatgc caggccagag gcagccctag
     1201 aagcaaccct gtcagactgc tgttcagcag cgacagcctg attctgcagg ccccttactc
     1261 cgtgttcgag ggcgatacac tggtgctgag atgccaccgg cggagaaaag aaaagctgac
     1321 cgccgtgaag tacacctgga acggcaacat cctgagcatc agcaacaaga gctgggacct
     1381 gctgatccct caggccagca gcaacaacaa cggcaactac cggtgtatcg gctacggcga
     1441 cgagaacgac gtgttcagaa gcaacttcaa gatcatcaag atccaagagc tgttcccgca
     1501 tccagagctg aaggccacag atagccagcc taccgagggc aactccgtga acctgtcttg
     1561 cgagacacag ctgcctccag agagaagcga tacccctctg cacttcaatt tcttccgcga
     1621 cggcgaagtg atcctgagcg attggagcac ataccccgaa ctgcagctgc ccaccgtttg
     1681 gagagagaac agcggcagct attggtgcgg agccgagaca gtgcggggca atatccacaa
     1741 gcacagcccc agcctgcaga tccacgtgca gagaattcca gtgtccggcg tgctgctgga
     1801 aacccaacct tctggtggac aggccgtgga aggcgaaatg ctggtgctcg tgtgttctgt
     1861 ggcagaaggc accggcgata ccaccttcag ctggcaccgg gaagatatgc aagagagcct
     1921 gggcagaaag acccagagaa gcctgagagc cgagctggaa ctgcctgcca tcagacagtc
     1981 tcacgctggc ggctactact gcaccgccga taatagctat ggccccgtgc agagcatggt
     2041 gctgaacgtg acagtcagag agactcccgg caatcgcgac ggacttggcg cgccggaaga
     2101 tcaagttgat ccaagattga ttgatggcaa gggcggatct ctggaggtgc tgttccaggg
     2161 accaggcgga agcggcaatc ccggcggcga tctggccccc cagatgctgc gcgagctgca
     2221 ggagaccaat gccgccctgc aggacgtgcg cgagctgctg cgccaccgcg tgaaggagat
     2281 caccttcctg aagaacaccg tgatggagtg cgacgcctgc ggtatgcagc cctctagagg
     2341 ctccggctcc atcatcccag ttgaggagga gaacccggac ttctggaacc gcgaggcagc
     2401 cgaggccctg ggtgccgcca agaagctgca gcctgcacag acagccgcca agaacctcat
     2461 catcttcctg ggcgatggga tgggggtgtc tacggtgaca gctgccagga tcctaaaagg
     2521 gcagaagaag gacaaactgg ggcctgagat acccctggcc atggaccgct tcccatatgt
     2581 ggctctgtcc aagacataca atgtagacaa acatgtgcca gacagtggag ccacagccac
     2641 ggcctacctg tgcggggtca agggcaactt ccagaccatt ggcttgagtg cagccgcccg
     2701 ctttaaccag tgcaacacga cacgcggcaa cgaggtcatc tccgtgatga atcgggccaa
     2761 gaaagcaggg aagtcagtgg gagtggtaac caccacacga gtgcagcacg cctcgccagc
     2821 cggcacctac gcccacacgg tgaaccgcaa ctggtactcg gacgccgacg tgcctgcctc
     2881 ggcccgccag gaggggtgcc aggacatcgc tacgcagctc atctccaaca tggacattga
     2941 cgtgatccta ggtggaggcc gaaagtacat gtttcgcatg ggaaccccag accctgagta
     3001 cccagatgac tacagccaag gtgggaccag gctggacggg aagaatctgg tgcaggaatg
     3061 gctggcgaag cgccagggtg cccggtatgt gtggaaccgc actgagctca tgcaggcttc
     3121 cctggacccg tctgtgaccc atctcatggg tctctttgag cctggagaca tgaaatacga
     3181 gatccaccga gactccacac tggacccctc cctgatggag atgacagagg ctgccctgcg
     3241 cctgctgagc aggaaccccc gcggcttctt cctcttcgtg gagggtggtc gcatcgacca
     3301 tggtcatcat gaaagcaggg cttaccgggc actgactgag acgatcatgt tcgacgacgc
     3361 cattgagagg gcgggccagc tcaccagcga ggaggacacg ctgagcctcg tcactgccga
     3421 ccactcccac gtcttctcct tcggaggcta ccccctgcga gggagctcca tcttcgggct
     3481 ggcccctggc aaggcccggg acaggaaggc ctacacggtc ctcctatacg gaaacggtcc
     3541 aggctatgtg ctcaaggacg gcgcccggcc ggatgttacc gagagcgaga gcgggagccc
     3601 cgagtatcgg cagcagtcag cagtgcccct ggacgaagag acccacgcag gcgaggacgt
     3661 ggcggtgttc gcgcgcggcc cgcaggcgca cctggttcac ggcgtgcagg agcagacctt
     3721 catagcgcac gtcatggcct tcgccgcctg cctggagccc tacaccgcct gcgacctggc
     3781 gccccccgcc ggcaccaccg acgccgcgca cccgggtggc ggaggactga acgacatctt
     3841 cgaggctcag aaaatcgaat ggcacgaaca tcaccatcac catcaccatc accattaagg
     3901 ccggcctgta gtcactggca caacgcgtta cccggtaagc caatcgggta tacacggtcg
     3961 tcatactgca gacagggttc ttctactttg caagatagtc ttgagtagta aaataaatag
     4021 atagagaaaa tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
     4081 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     4141 ggaaattgca tcacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     4201 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     4261 taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     4321 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     4381 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     4441 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     4501 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc
     4561 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt
     4621 gtatatccat tttcggatct gatcagcacg tgttgacaat taatcatcgg catagtatat
     4681 cggcatagta taatacgaca aggtgaggaa ctaaaccatg accgagtaca agcccacggt
     4741 gcgcctcgcc acccgcgacg acgtccccag ggccgtacgc accctcgccg ccgcgttcgc
     4801 cgactacccc gccacgcgcc acaccgtcga tccggaccgc cacatcgagc gggtcaccga
     4861 gctgcaagaa ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga
     4921 cgacggcgcc gcggtggcgg tctggaccac gccggagagc gtcgaagcgg gggcggtgtt
     4981 cgccgagatc ggcccgcgca tggccgagtt gagcggttcc cggctggccg cgcagcaaca
     5041 gatggaaggc ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc tggccaccgt
     5101 cggcgtctcg cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc tccccggagt
     5161 ggaggcggcc gagcgcgccg gggtgcccgc cttcctggag acatccgcgc cccgcaacct
     5221 ccccttctac gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc
     5281 gcgcacctgg tgcatgaccc gcaagcccgg tgcctgacac gtgctacgag atttcgattc
     5341 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     5401 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc
     5461 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     5521 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat
     5581 accgtcgacc tctagctaga gcttggcgta atcatggtca ttaccaatgc ttaatcagtg
     5641 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     5701 tgtagataac tacgatacgg gagggcttac catctggccc cagcgctgcg atgataccgc
     5761 gagaaccacg ctcaccggct ccggatttat cagcaataaa ccagccagcc ggaagggccg
     5821 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     5881 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc atcgctacag
     5941 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     6001 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     6061 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     6121 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     6181 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     6241 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     6301 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     6361 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     6421 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     6481 tattcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     6541 acatatttga atgtatttag aaaaataaac aaataggggt cagtgttaca accaattaac
     6601 caattctgaa cattatcgcg agcccattta tacctgaata tggctcataa caccccttgc
     6661 tcatgaccaa aatcccttaa cgtgagttac gcgcgcgtcg ttccactgag cgtcagaccc
     6721 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     6781 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     6841 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     6901 gtagccgtag ttagcccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     6961 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     7021 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     7081 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     7141 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     7201 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     7261 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     7321 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     7381 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     7441 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     7501 cgaggaagcg gaaggcgaga gtagggaact gccaggcatc aaactaagca gaaggcccct
     7561 gacggatggc ctttttgcgt ttctacaaac tctttctgtg ttgtaaaacg acggccagtc
     7621 ttaagctcgg gccccctggg cggttctgat aacgagtaat cgttaatccg caaataacgt
     7681 aaaaacccgc ttcggcgggt ttttttatgg ggggagttta gggaaagagc atttgtcaga
     7741 atatttaagg gcgcctgtca ctttgcttga tatatgagaa ttatttaacc ttataaatga
     7801 gaaaaaagca acgcacttta aataagatac gttgcttttt cgattgatga acacctataa
     7861 ttaaactatt catctattat ttatgatttt ttgtatatac aatatttcta gtttgttaaa
     7921 gagaattaag aaaataaatc tcgaaaataa taaagggaaa atcagttttt gatatcaaaa
     7981 ttatacatgt caacgataat acaaaatata atacaaacta taagatgtta tcagtattta
     8041 ttatcattta gaataaattt tgtgtcgccc ttaattgtga gcggataaca attacgagct
     8101 tcatgcacag tggcgttgac tcaatattgg ccattagcca tattattcat tggttatata
     8161 gcataaatca atattggcta ttggccattg catacgttgt atctatatca taatatgtac
     8221 atttatattg gctcatgtcc aatatgaccg ccatgtt
//
