LOCUS       sequence-460930-        7960 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 7960)
  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 7960)
  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-SPACA6-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..7960
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     91..108
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(248..274)
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     CDS             complement(275..319)
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             complement(1979..2002)
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             complement(2012..2047)
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     primer_bind     complement(2981..3000)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        complement(3013..3031)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3014..3031)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3241..3265)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        complement(3262..3465)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(3291..3311)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        3466..3845
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     protein_bind    4012..4028
                     /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)."
     primer_bind     complement(4484..4501)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4484..4500)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4599..4616)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4770..5358)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4850..4869)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(5444..5463)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(5469..5488)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        5530..5621
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5622..5690,5691..6482)
                     /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     complement(5840..5859)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     polyA_signal    6535..6656
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6546..6565)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6600..6619
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(6786..7385)
                     /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(6869..6889)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     7366..7385
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(7404..7451)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(7499..7828)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7513..7648
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(7567..7586)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     7813..7833
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    join(7951..7960,1..102)
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
ORIGIN
        1 cctcatttta ttaggaaagg acagtgggag tggcaccttc cagggtcaag gaaggcacgg
       61 gggaggggca aacaacagat ggctggcaac tagaaggcac agtcgaggct gattttctct
      121 atctatttat tttactactc aagactatct tgcaaagtag aagaaccctg tctgcagtat
      181 gacgaccgtg tatacccgat tggcttaccg ggtaacgcgt tgtgccagtg actacaggcc
      241 ggccttaatg gtgatggtga tggtgatggt gatgttcgtg ccattcgatt ttctgagcct
      301 cgaagatgtc gttcagtcct ccgccacccg ggtgcgcggc gtcggtggtg ccggcggggg
      361 gcgccaggtc gcaggcggtg tagggctcca ggcaggcggc gaaggccatg acgtgcgcta
      421 tgaaggtctg ctcctgcacg ccgtgaacca ggtgcgcctg cgggccgcgc gcgaacaccg
      481 ccacgtcctc gcctgcgtgg gtctcttcgt ccaggggcac tgctgactgc tgccgatact
      541 cggggctccc gctctcgctc tcggtaacat ccggccgggc gccgtccttg agcacatagc
      601 ctggaccgtt tccgtatagg aggaccgtgt aggccttcct gtcccgggcc ttgccagggg
      661 ccagcccgaa gatggagctc cctcgcaggg ggtagcctcc gaaggagaag acgtgggagt
      721 ggtcggcagt gacgaggctc agcgtgtcct cctcgctggt gagctggccc gccctctcaa
      781 tggcgtcgtc gaacatgatc gtctcagtca gtgcccggta agccctgctt tcatgatgac
      841 catggtcgat gcgaccaccc tccacgaaga ggaagaagcc gcgggggttc ctgctcagca
      901 ggcgcagggc agcctctgtc atctccatca gggaggggtc cagtgtggag tctcggtgga
      961 tctcgtattt catgtctcca ggctcaaaga gacccatgag atgggtcaca gacgggtcca
     1021 gggaagcctg catgagctca gtgcggttcc acacataccg ggcaccctgg cgcttcgcca
     1081 gccattcctg caccagattc ttcccgtcca gcctggtccc accttggctg tagtcatctg
     1141 ggtactcagg gtctggggtt cccatgcgaa acatgtactt tcggcctcca cctaggatca
     1201 cgtcaatgtc catgttggag atgagctgcg tagcgatgtc ctggcacccc tcctggcggg
     1261 ccgaggcagg cacgtcggcg tccgagtacc agttgcggtt caccgtgtgg gcgtaggtgc
     1321 cggctggcga ggcgtgctgc actcgtgtgg tggttaccac tcccactgac ttccctgctt
     1381 tcttggcccg attcatcacg gagatgacct cgttgccgcg tgtcgtgttg cactggttaa
     1441 agcgggcggc tgcactcaag ccaatggtct ggaagttgcc cttgaccccg cacaggtagg
     1501 ccgtggctgt ggctccactg tctggcacat gtttgtctac attgtatgtc ttggacagag
     1561 ccacatatgg gaagcggtcc atggccaggg gtatctcagg ccccagtttg tccttcttct
     1621 gcccttttag gatcctggca gctgtcaccg tagacacccc catcccatcg cccaggaaga
     1681 tgatgaggtt cttggcggct gtctgtgcag gctgcagctt cttggcggca cccagggcct
     1741 cggctgcctc gcggttccag aagtccgggt tctcctcctc aactgggatg atggagccgg
     1801 agcctctaga gggctgcata ccgcaggcgt cgcactccat cacggtgttc ttcaggaagg
     1861 tgatctcctt cacgcggtgg cgcagcagct cgcgcacgtc ctgcagggcg gcattggtct
     1921 cctgcagctc gcgcagcatc tggggggcca gatcgccgcc gggattgccg cttccgcctg
     1981 gtccctggaa cagcacctcc agagatccgc ccttgccatc aatcaatctt ggatcaactt
     2041 gatcttccgg cgcgccaagg ttgctagggg tcagagcctc aggtctagca agcagctctc
     2101 ccagagaagg cctccaaggc tcaatcagct cggcatctct aggggcccat ctaagcactt
     2161 cccggaagct ggcctgcagt tctgtttcgg ctcttggagg aggtccggtc acattcagga
     2221 agaagtacag tctagccagg ggccgctgat cctgcttgat cacgcaactg aaggtgcctc
     2281 tgtgggtcag ctgagcaggt ctaattctgg ccaggtagcc ctcggctctg ggcatgtctc
     2341 tgaagtagct caggtcctgg gttctcaggc ctccgccggc aaacttccaa gagtaggtga
     2401 tttcctcttt aggcagctgg aagttcacga tgcagctgaa catggcctgg tcgcctcttg
     2461 tcactgtcac atcctgcaca gggcagtcca gaggcagatc gcacactctg ctgtagcagc
     2521 cgctgcacag aaaccgtctg gcgaactctt gaaggccgca tggaggaatg caggcctggg
     2581 cctctttcag ctgggtgatc actttcttca tcttctcggc ggcgtcagga aatgccacct
     2641 caaagcttcc ctgagcagcg gccagttctt gcagggcgtg tgtcatctgt gtgaaggtgt
     2701 cgtgcagatg gcttctctcg tcgtagttga tctcggtgtc gctcaggccc tgaaaggcgg
     2761 ctgtaaaggc ttcctcgcac tcttccagct tggggcttct catgcccaca aacatctggc
     2821 agatccgcag ccgctcgctg taggtggtaa aacacagcag acaggcggcc gcagcgaaca
     2881 ccaggcagaa gatgtagctc agggcaatga tggtcttcat ggtggcgcta gcaggcgcgt
     2941 tttcgataga accgaggtgc agttggacct gggagtggac acctgtggag agaaaggcaa
     3001 agtggatgtt attctatagt gtcacctaaa tcgtatgtgt atgatacata aggttatgta
     3061 ttaattgtag ccgcgttcta acgaagccaa gggggtgggc ctatagactc tataggcggt
     3121 acttacgtca ctcttggcac ggggaatccg cgttccaatg caccgttccc ggccgcggag
     3181 gctggatcgg tcccggtgtc ttctatggag gtcaaaacag cgtggatggc gtctccaggc
     3241 gatctgacgg ttcactaaac gagctctgct tatatagacc tcccaccgta cacgcctacc
     3301 gcccatttgc gtcaatgggg cggagttgtt acgacatttt ggaaagtccc gttgattttg
     3361 gtgccaaaac aaactcccat tgacgtcaat ggggtggaga cttggaaatc cccgtgagtc
     3421 aaaccgctat ccacgcccat tgatgtactg ccaaaaccgc atcaccatgg taatagcgat
     3481 gactaatacg tagatgtact gccaagtagg aaagtcccat aaggtcatgt actgggcata
     3541 atgccaggcg ggccatttac cgtcattgac gtcaataggg ggcgtacttg gcatatgata
     3601 cacttgatgt actgccaagt gggcagttta ccgtaaatac tccacccatt gacgtcaatg
     3661 gaaagtccct attggcgtta ctatgggaac atacgtcatt attgacgtca atgggcgggg
     3721 gtcgttgggc ggtcagccag gcgggccatt taccgtaagt tatgtaacgc ggaactccat
     3781 atatgggcta tgaactaatg accccgtaat tgattactat taataactag tcaataatca
     3841 atgccaacat ggcggtcata ttggacatga gccaatataa atgtacatat tatgatatag
     3901 atacaacgta tgcaatggcc aatagccaat attgatttat gctatataac caatgaataa
     3961 tatggctaat ggccaatatt gagtcaacgc cactgtgcat gaagctcgta attgttatcc
     4021 gctcacaatt aagggcgaca caaaatttat tctaaatgat aataaatact gataacatct
     4081 tatagtttgt attatatttt gtattatcgt tgacatgtat aattttgata tcaaaaactg
     4141 attttccctt tattattttc gagatttatt ttcttaattc tctttaacaa actagaaata
     4201 ttgtatatac aaaaaatcat aaataataga tgaatagttt aattataggt gttcatcaat
     4261 cgaaaaagca acgtatctta tttaaagtgc gttgcttttt tctcatttat aaggttaaat
     4321 aattctcata tatcaagcaa agtgacaggc gcccttaaat attctgacaa atgctctttc
     4381 cctaaactcc ccccataaaa aaacccgccg aagcgggttt ttacgttatt tgcggattaa
     4441 cgattactcg ttatcagaac cgcccagggg gcccgagctt aagactggcc gtcgttttac
     4501 aacacagaaa gagtttgtag aaacgcaaaa aggccatccg tcaggggcct tctgcttagt
     4561 ttgatgcctg gcagttccct actctcgcct tccgcttcct cgctcactga ctcgctgcgc
     4621 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     4681 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     4741 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     4801 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     4861 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     4921 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     4981 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     5041 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     5101 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     5161 ggtgctacag agttcttgaa gtggtgggct aactacggct acactagaag aacagtattt
     5221 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     5281 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     5341 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     5401 aacgacgcgc gcgtaactca cgttaaggga ttttggtcat gagcaagggg tgttatgagc
     5461 catattcagg tataaatggg ctcgcgataa tgttcagaat tggttaattg gttgtaacac
     5521 tgacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     5581 aaccctgata aatgcttcaa taatattgaa aaaggaagaa tatgagtatt caacatttcc
     5641 gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa
     5701 cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac
     5761 tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga
     5821 tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag
     5881 agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca
     5941 cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca
     6001 tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa
     6061 ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc
     6121 tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagcg atggcaacaa
     6181 cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag
     6241 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct
     6301 ggtttattgc tgataaatcc ggagccggtg agcgtggttc tcgcggtatc atcgcagcgc
     6361 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa
     6421 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt
     6481 aatgaccatg attacgccaa gctctagcta gaggtcgacg gtatacagac atgataagat
     6541 acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg
     6601 aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttgggg
     6661 tgggcgaaga actccagcat gagatccccg cgctggagga tcatccagcc ggcgtcccgg
     6721 aaaacgattc cgaagcccaa cctttcatag aaggcggcgg tggaatcgaa atctcgtagc
     6781 acgtgtcagg caccgggctt gcgggtcatg caccaggtgc gcggtccttc gggcacctcg
     6841 acgtcggcgg tgacggtgaa gccgagccgc tcgtagaagg ggaggttgcg gggcgcggat
     6901 gtctccagga aggcgggcac cccggcgcgc tcggccgcct ccactccggg gagcacgacg
     6961 gcgctgccca gacccttgcc ctggtggtcg ggcgagacgc cgacggtggc caggaaccac
     7021 gcgggctcct tgggccggtg cggcgccagg aggccttcca tctgttgctg cgcggccagc
     7081 cgggaaccgc tcaactcggc catgcgcggg ccgatctcgg cgaacaccgc ccccgcttcg
     7141 acgctctccg gcgtggtcca gaccgccacc gcggcgccgt cgtccgcgac ccacaccttg
     7201 ccgatgtcga gcccgacgcg cgtgaggaag agttcttgca gctcggtgac ccgctcgatg
     7261 tggcggtccg gatcgacggt gtggcgcgtg gcggggtagt cggcgaacgc ggcggcgagg
     7321 gtgcgtacgg ccctggggac gtcgtcgcgg gtggcgaggc gcaccgtggg cttgtactcg
     7381 gtcatggttt agttcctcac cttgtcgtat tatactatgc cgatatacta tgccgatgat
     7441 taattgtcaa cacgtgctga tcagatccga aaatggatat acaagctccc gggagctttt
     7501 tgcaaaagcc taggcctcca aaaaagcctc ctcactactt ctggaatagc tcagaggcag
     7561 aggcggcctc ggcctctgca taaataaaaa aaattagtca gccatggggc ggagaatggg
     7621 cggaactggg cggagttagg ggcgggatgg gcggagttag gggcgggact atggttgctg
     7681 actaattgag atgcatgctt tgcatacttc tgcctgctgg ggagcctggg gactttccac
     7741 acctggttgc tgactaattg agatgcatgc tttgcatact tctgcctgct ggggagcctg
     7801 gggactttcc acaccctaac tgacacacat tccacagaat taattcgcgt taaatttttg
     7861 ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa
     7921 agaatagacc gagatagggt tgagtgttgt gatgcaattt
//
