LOCUS       sequence-401012-        7687 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 7687)
  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 7687)
  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-PILRA-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..7687
                     /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             1526..1561
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             1571..1594
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             3254..3298
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             3299..3325
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(3465..3482)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3471..3582
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3700..3720)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3705..4034
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3885..4020
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3947..3966
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4082..4129
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4148..4747
                     /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(4148..4167)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4644..4664
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4877..4998
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4914..4933)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4968..4987
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(5051..5842,5843..5911))
                     /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     5674..5693
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5912..6003)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6045..6064
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6070..6089
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      6175..6763
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6664..6683
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6917..6934
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     7032..7049
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7033..7049
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    7505..7521
                     /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 cccaaccttc tggatctact ggctctggcc ctagctacct
     1021 gtatggcgtg acacagccta agcacctgtc tgcctctatg ggcggcagcg tggaaatccc
     1081 cttcagcttc tactacccct gggaactcgc cacagctcct gatgtgcgga tctcttggag
     1141 aagaggccac ttccaccggc agagcttcta cagcacaaga ccacctagca tccacaagga
     1201 ctacgtgaac cggctgttcc tgaactggac cgagggccag aagtccggct tcctgagaat
     1261 cagcaacctg cagaaacagg accagagcgt gtacttctgc agagtggaac tggacaccag
     1321 aagcagcggt agacagcagt ggcagagcat cgagggcacc aagctgtcta tcacccaggc
     1381 cgtgaccacc acaacacaga ggcctagcag catgaccaca acatggcggc tgagcagcac
     1441 cacaaccaca acaggcctga gagtgaccca gggcaagaga agaagcgaca gctggcacat
     1501 cagcctggaa accgctggcg cgccggaaga tcaagttgat ccaagattga ttgatggcaa
     1561 gggcggatct ctggaggtgc tgttccaggg accaggcgga agcggcaatc ccggcggcga
     1621 tctggccccc cagatgctgc gcgagctgca ggagaccaat gccgccctgc aggacgtgcg
     1681 cgagctgctg cgccaccgcg tgaaggagat caccttcctg aagaacaccg tgatggagtg
     1741 cgacgcctgc ggtatgcagc cctctagagg ctccggctcc atcatcccag ttgaggagga
     1801 gaacccggac ttctggaacc gcgaggcagc cgaggccctg ggtgccgcca agaagctgca
     1861 gcctgcacag acagccgcca agaacctcat catcttcctg ggcgatggga tgggggtgtc
     1921 tacggtgaca gctgccagga tcctaaaagg gcagaagaag gacaaactgg ggcctgagat
     1981 acccctggcc atggaccgct tcccatatgt ggctctgtcc aagacataca atgtagacaa
     2041 acatgtgcca gacagtggag ccacagccac ggcctacctg tgcggggtca agggcaactt
     2101 ccagaccatt ggcttgagtg cagccgcccg ctttaaccag tgcaacacga cacgcggcaa
     2161 cgaggtcatc tccgtgatga atcgggccaa gaaagcaggg aagtcagtgg gagtggtaac
     2221 caccacacga gtgcagcacg cctcgccagc cggcacctac gcccacacgg tgaaccgcaa
     2281 ctggtactcg gacgccgacg tgcctgcctc ggcccgccag gaggggtgcc aggacatcgc
     2341 tacgcagctc atctccaaca tggacattga cgtgatccta ggtggaggcc gaaagtacat
     2401 gtttcgcatg ggaaccccag accctgagta cccagatgac tacagccaag gtgggaccag
     2461 gctggacggg aagaatctgg tgcaggaatg gctggcgaag cgccagggtg cccggtatgt
     2521 gtggaaccgc actgagctca tgcaggcttc cctggacccg tctgtgaccc atctcatggg
     2581 tctctttgag cctggagaca tgaaatacga gatccaccga gactccacac tggacccctc
     2641 cctgatggag atgacagagg ctgccctgcg cctgctgagc aggaaccccc gcggcttctt
     2701 cctcttcgtg gagggtggtc gcatcgacca tggtcatcat gaaagcaggg cttaccgggc
     2761 actgactgag acgatcatgt tcgacgacgc cattgagagg gcgggccagc tcaccagcga
     2821 ggaggacacg ctgagcctcg tcactgccga ccactcccac gtcttctcct tcggaggcta
     2881 ccccctgcga gggagctcca tcttcgggct ggcccctggc aaggcccggg acaggaaggc
     2941 ctacacggtc ctcctatacg gaaacggtcc aggctatgtg ctcaaggacg gcgcccggcc
     3001 ggatgttacc gagagcgaga gcgggagccc cgagtatcgg cagcagtcag cagtgcccct
     3061 ggacgaagag acccacgcag gcgaggacgt ggcggtgttc gcgcgcggcc cgcaggcgca
     3121 cctggttcac ggcgtgcagg agcagacctt catagcgcac gtcatggcct tcgccgcctg
     3181 cctggagccc tacaccgcct gcgacctggc gccccccgcc ggcaccaccg acgccgcgca
     3241 cccgggtggc ggaggactga acgacatctt cgaggctcag aaaatcgaat ggcacgaaca
     3301 tcaccatcac catcaccatc accattaagg ccggcctgta gtcactggca caacgcgtta
     3361 cccggtaagc caatcgggta tacacggtcg tcatactgca gacagggttc ttctactttg
     3421 caagatagtc ttgagtagta aaataaatag atagagaaaa tcagcctcga ctgtgccttc
     3481 tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc
     3541 cactcccact gtcctttcct aataaaatga ggaaattgca tcacaacact caaccctatc
     3601 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     3661 gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt
     3721 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     3781 cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     3841 atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc
     3901 cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg
     3961 ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
     4021 taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct gatcagcacg
     4081 tgttgacaat taatcatcgg catagtatat cggcatagta taatacgaca aggtgaggaa
     4141 ctaaaccatg accgagtaca agcccacggt gcgcctcgcc acccgcgacg acgtccccag
     4201 ggccgtacgc accctcgccg ccgcgttcgc cgactacccc gccacgcgcc acaccgtcga
     4261 tccggaccgc cacatcgagc gggtcaccga gctgcaagaa ctcttcctca cgcgcgtcgg
     4321 gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg tctggaccac
     4381 gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca tggccgagtt
     4441 gagcggttcc cggctggccg cgcagcaaca gatggaaggc ctcctggcgc cgcaccggcc
     4501 caaggagccc gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc agggcaaggg
     4561 tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg gggtgcccgc
     4621 cttcctggag acatccgcgc cccgcaacct ccccttctac gagcggctcg gcttcaccgt
     4681 caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc gcaagcccgg
     4741 tgcctgacac gtgctacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
     4801 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     4861 gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     4921 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     4981 actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta
     5041 atcatggtca ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     5101 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     5161 catctggccc cagcgctgcg atgataccgc gagaaccacg ctcaccggct ccggatttat
     5221 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     5281 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     5341 gtttgcgcaa cgttgttgcc atcgctacag gcatcgtggt gtcacgctcg tcgtttggta
     5401 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     5461 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     5521 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     5581 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     5641 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     5701 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     5761 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     5821 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     5881 taagggcgac acggaaatgt tgaatactca tattcttcct ttttcaatat tattgaagca
     5941 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     6001 aaataggggt cagtgttaca accaattaac caattctgaa cattatcgcg agcccattta
     6061 tacctgaata tggctcataa caccccttgc tcatgaccaa aatcccttaa cgtgagttac
     6121 gcgcgcgtcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     6181 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     6241 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     6301 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttagcccacc acttcaagaa
     6361 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     6421 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     6481 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     6541 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     6601 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     6661 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     6721 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     6781 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     6841 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     6901 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaaggcgaga gtagggaact
     6961 gccaggcatc aaactaagca gaaggcccct gacggatggc ctttttgcgt ttctacaaac
     7021 tctttctgtg ttgtaaaacg acggccagtc ttaagctcgg gccccctggg cggttctgat
     7081 aacgagtaat cgttaatccg caaataacgt aaaaacccgc ttcggcgggt ttttttatgg
     7141 ggggagttta gggaaagagc atttgtcaga atatttaagg gcgcctgtca ctttgcttga
     7201 tatatgagaa ttatttaacc ttataaatga gaaaaaagca acgcacttta aataagatac
     7261 gttgcttttt cgattgatga acacctataa ttaaactatt catctattat ttatgatttt
     7321 ttgtatatac aatatttcta gtttgttaaa gagaattaag aaaataaatc tcgaaaataa
     7381 taaagggaaa atcagttttt gatatcaaaa ttatacatgt caacgataat acaaaatata
     7441 atacaaacta taagatgtta tcagtattta ttatcattta gaataaattt tgtgtcgccc
     7501 ttaattgtga gcggataaca attacgagct tcatgcacag tggcgttgac tcaatattgg
     7561 ccattagcca tattattcat tggttatata gcataaatca atattggcta ttggccattg
     7621 catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc aatatgaccg
     7681 ccatgtt
//
