LOCUS       sequence-421009-        7822 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 7822)
  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 7822)
  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-NRG1-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..7822
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(27..118)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     160..179
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     185..204
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      290..878
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     779..798
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1032..1049
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     1147..1164
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1148..1164
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1620..1636
                     /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        1803..2182
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2183..2386
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2337..2357
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2383..2407
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        2617..2635
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2617..2634
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     2648..2667
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             3463..3498
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             3508..3531
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             5191..5235
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5236..5262
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(5402..5419)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5408..5519
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5637..5657)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5642..5971
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5822..5957
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5884..5903
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6019..6066
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6085..6684
                     /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(6085..6104)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6581..6601
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6814..6935
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6851..6870)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6905..6924
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(6988..7779,7780..7822,1..26))
                     /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     7611..7630
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 gcgacacgga aatgttgaat actcatattc ttcctttttc aatattattg aagcatttat
       61 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
      121 ggggtcagtg ttacaaccaa ttaaccaatt ctgaacatta tcgcgagccc atttatacct
      181 gaatatggct cataacaccc cttgctcatg accaaaatcc cttaacgtga gttacgcgcg
      241 cgtcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
      301 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
      361 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
      421 agataccaaa tactgttctt ctagtgtagc cgtagttagc ccaccacttc aagaactctg
      481 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
      541 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
      601 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
      661 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
      721 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
      781 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
      841 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
      901 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
      961 attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa
     1021 cgaccgagcg cagcgagtca gtgagcgagg aagcggaagg cgagagtagg gaactgccag
     1081 gcatcaaact aagcagaagg cccctgacgg atggcctttt tgcgtttcta caaactcttt
     1141 ctgtgttgta aaacgacggc cagtcttaag ctcgggcccc ctgggcggtt ctgataacga
     1201 gtaatcgtta atccgcaaat aacgtaaaaa cccgcttcgg cgggtttttt tatgggggga
     1261 gtttagggaa agagcatttg tcagaatatt taagggcgcc tgtcactttg cttgatatat
     1321 gagaattatt taaccttata aatgagaaaa aagcaacgca ctttaaataa gatacgttgc
     1381 tttttcgatt gatgaacacc tataattaaa ctattcatct attatttatg attttttgta
     1441 tatacaatat ttctagtttg ttaaagagaa ttaagaaaat aaatctcgaa aataataaag
     1501 ggaaaatcag tttttgatat caaaattata catgtcaacg ataatacaaa atataataca
     1561 aactataaga tgttatcagt atttattatc atttagaata aattttgtgt cgcccttaat
     1621 tgtgagcgga taacaattac gagcttcatg cacagtggcg ttgactcaat attggccatt
     1681 agccatatta ttcattggtt atatagcata aatcaatatt ggctattggc cattgcatac
     1741 gttgtatcta tatcataata tgtacattta tattggctca tgtccaatat gaccgccatg
     1801 ttggcattga ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag
     1861 cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
     1921 caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg
     1981 gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca
     2041 tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc
     2101 ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt
     2161 attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata
     2221 gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt
     2281 ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca
     2341 aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctcgtt tagtgaaccg
     2401 tcagatcgcc tggagacgcc atccacgctg ttttgacctc catagaagac accgggaccg
     2461 atccagcctc cgcggccggg aacggtgcat tggaacgcgg attccccgtg ccaagagtga
     2521 cgtaagtacc gcctatagag tctataggcc cacccccttg gcttcgttag aacgcggcta
     2581 caattaatac ataaccttat gtatcataca catacgattt aggtgacact atagaataac
     2641 atccactttg cctttctctc cacaggtgtc cactcccagg tccaactgca cctcggttct
     2701 atcgaaaacg cgcctgctag cgccaccatg aagaccatca ttgccctgag ctacatcttc
     2761 tgcctggtgt tcgctgcggc cgcctctgga aagaaacctg aatctgccgc cggatctcag
     2821 agccctgcac tgcctcctag actgaaagag atgaagtccc aagagagcgc cgctggctct
     2881 aagctggtgc tgagatgtga aaccagcagc gagtacagca gcctgcggtt caagtggttc
     2941 aagaacggca acgagctgaa ccggaagaac aagccccaga acatcaagat ccagaagaag
     3001 cccggcaaga gcgagctgag aatcaacaag gcctctctgg ccgactccgg cgagtacatg
     3061 tgcaaagtga tcagcaagct gggcaacgac agcgccagcg ccaatatcac catcgtggaa
     3121 agcaacgaga tcatcaccgg catgcctgcc tctacagagg gcgcctatgt gtctagcgag
     3181 agccccatca gaatcagcgt gtccacagaa ggcgccaaca ccagcagcag cacaagcacc
     3241 tctacaacag gcaccagcca cctggtcaag tgcgccgaga aagaaaagac cttctgcgtg
     3301 aacggcggcg agtgcttcat ggtcaaggac ctgagcaacc ccagcagata cctgtgcaag
     3361 tgccagcctg gctttaccgg cgccagatgt accgagaatg tgcccatgaa ggtgcagaat
     3421 caagagaagg ccgaggaact gtaccagaaa cgaggcgcgc cggaagatca agttgatcca
     3481 agattgattg atggcaaggg cggatctctg gaggtgctgt tccagggacc aggcggaagc
     3541 ggcaatcccg gcggcgatct ggccccccag atgctgcgcg agctgcagga gaccaatgcc
     3601 gccctgcagg acgtgcgcga gctgctgcgc caccgcgtga aggagatcac cttcctgaag
     3661 aacaccgtga tggagtgcga cgcctgcggt atgcagccct ctagaggctc cggctccatc
     3721 atcccagttg aggaggagaa cccggacttc tggaaccgcg aggcagccga ggccctgggt
     3781 gccgccaaga agctgcagcc tgcacagaca gccgccaaga acctcatcat cttcctgggc
     3841 gatgggatgg gggtgtctac ggtgacagct gccaggatcc taaaagggca gaagaaggac
     3901 aaactggggc ctgagatacc cctggccatg gaccgcttcc catatgtggc tctgtccaag
     3961 acatacaatg tagacaaaca tgtgccagac agtggagcca cagccacggc ctacctgtgc
     4021 ggggtcaagg gcaacttcca gaccattggc ttgagtgcag ccgcccgctt taaccagtgc
     4081 aacacgacac gcggcaacga ggtcatctcc gtgatgaatc gggccaagaa agcagggaag
     4141 tcagtgggag tggtaaccac cacacgagtg cagcacgcct cgccagccgg cacctacgcc
     4201 cacacggtga accgcaactg gtactcggac gccgacgtgc ctgcctcggc ccgccaggag
     4261 gggtgccagg acatcgctac gcagctcatc tccaacatgg acattgacgt gatcctaggt
     4321 ggaggccgaa agtacatgtt tcgcatggga accccagacc ctgagtaccc agatgactac
     4381 agccaaggtg ggaccaggct ggacgggaag aatctggtgc aggaatggct ggcgaagcgc
     4441 cagggtgccc ggtatgtgtg gaaccgcact gagctcatgc aggcttccct ggacccgtct
     4501 gtgacccatc tcatgggtct ctttgagcct ggagacatga aatacgagat ccaccgagac
     4561 tccacactgg acccctccct gatggagatg acagaggctg ccctgcgcct gctgagcagg
     4621 aacccccgcg gcttcttcct cttcgtggag ggtggtcgca tcgaccatgg tcatcatgaa
     4681 agcagggctt accgggcact gactgagacg atcatgttcg acgacgccat tgagagggcg
     4741 ggccagctca ccagcgagga ggacacgctg agcctcgtca ctgccgacca ctcccacgtc
     4801 ttctccttcg gaggctaccc cctgcgaggg agctccatct tcgggctggc ccctggcaag
     4861 gcccgggaca ggaaggccta cacggtcctc ctatacggaa acggtccagg ctatgtgctc
     4921 aaggacggcg cccggccgga tgttaccgag agcgagagcg ggagccccga gtatcggcag
     4981 cagtcagcag tgcccctgga cgaagagacc cacgcaggcg aggacgtggc ggtgttcgcg
     5041 cgcggcccgc aggcgcacct ggttcacggc gtgcaggagc agaccttcat agcgcacgtc
     5101 atggccttcg ccgcctgcct ggagccctac accgcctgcg acctggcgcc ccccgccggc
     5161 accaccgacg ccgcgcaccc gggtggcgga ggactgaacg acatcttcga ggctcagaaa
     5221 atcgaatggc acgaacatca ccatcaccat caccatcacc attaaggccg gcctgtagtc
     5281 actggcacaa cgcgttaccc ggtaagccaa tcgggtatac acggtcgtca tactgcagac
     5341 agggttcttc tactttgcaa gatagtcttg agtagtaaaa taaatagata gagaaaatca
     5401 gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc
     5461 ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatca
     5521 caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg
     5581 cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa
     5641 tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     5701 catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag
     5761 aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc
     5821 catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
     5881 ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga agtagtgagg
     5941 aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta tatccatttt
     6001 cggatctgat cagcacgtgt tgacaattaa tcatcggcat agtatatcgg catagtataa
     6061 tacgacaagg tgaggaacta aaccatgacc gagtacaagc ccacggtgcg cctcgccacc
     6121 cgcgacgacg tccccagggc cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc
     6181 acgcgccaca ccgtcgatcc ggaccgccac atcgagcggg tcaccgagct gcaagaactc
     6241 ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg
     6301 gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc
     6361 ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc agcaacagat ggaaggcctc
     6421 ctggcgccgc accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc
     6481 gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag
     6541 cgcgccgggg tgcccgcctt cctggagaca tccgcgcccc gcaacctccc cttctacgag
     6601 cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc
     6661 atgacccgca agcccggtgc ctgacacgtg ctacgagatt tcgattccac cgccgccttc
     6721 tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc
     6781 ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt
     6841 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     6901 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct
     6961 agctagagct tggcgtaatc atggtcatta ccaatgctta atcagtgagg cacctatctc
     7021 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     7081 gatacgggag ggcttaccat ctggccccag cgctgcgatg ataccgcgag aaccacgctc
     7141 accggctccg gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     7201 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     7261 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatc gctacaggca tcgtggtgtc
     7321 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     7381 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     7441 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     7501 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     7561 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     7621 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     7681 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     7741 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     7801 tgccgcaaaa aagggaataa gg
//
