LOCUS       sequence-424380-        7693 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 7693)
  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 7693)
  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-FCGR3B-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..7693
                     /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             1532..1567
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             1577..1600
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             3260..3304
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             3305..3331
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(3471..3488)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3477..3588
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3706..3726)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3711..4040
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3891..4026
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3953..3972
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4088..4135
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4154..4753
                     /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(4154..4173)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4650..4670
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4883..5004
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4920..4939)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4974..4993
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(5057..5848,5849..5917))
                     /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     5680..5699
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5918..6009)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6051..6070
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6076..6095
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      6181..6769
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6670..6689
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6923..6940
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     7038..7055
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7039..7055
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    7511..7527
                     /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 ccatgagaac agaggatctg cctaaagccg tggtgttcct
     1021 ggaacctcag tggtacagcg tgctggaaaa ggacagcgtg accctgaagt gtcagggcgc
     1081 ctactctccc gaggataaca gcacccagtg gttccacaac gagagcctga tcagcagcca
     1141 ggccagcagc tactttatcg acgccgccac agtgaacgac agcggcgagt acagatgcca
     1201 gaccaatctg agcaccctga gcgaccctgt gcagctggaa gtgcatattg gatggctgct
     1261 gctgcaggcc cctagatggg tgttcaaaga agaggacccc atccacctga gatgccactc
     1321 ttggaagaac acagccctgc acaaagtgac ctacctgcag aacggcaagg acagaaagta
     1381 cttccaccac aacagcgact ttcacatccc caaggccaca ctgaaggact ccggctccta
     1441 tttctgtaga ggcctcgtgg gcagcaagaa cgtgtccagc gagacagtga acatcaccat
     1501 cacacagggc ctcgccgtgt ctggcgcgcc ggaagatcaa gttgatccaa gattgattga
     1561 tggcaagggc ggatctctgg aggtgctgtt ccagggacca ggcggaagcg gcaatcccgg
     1621 cggcgatctg gccccccaga tgctgcgcga gctgcaggag accaatgccg ccctgcagga
     1681 cgtgcgcgag ctgctgcgcc accgcgtgaa ggagatcacc ttcctgaaga acaccgtgat
     1741 ggagtgcgac gcctgcggta tgcagccctc tagaggctcc ggctccatca tcccagttga
     1801 ggaggagaac ccggacttct ggaaccgcga ggcagccgag gccctgggtg ccgccaagaa
     1861 gctgcagcct gcacagacag ccgccaagaa cctcatcatc ttcctgggcg atgggatggg
     1921 ggtgtctacg gtgacagctg ccaggatcct aaaagggcag aagaaggaca aactggggcc
     1981 tgagataccc ctggccatgg accgcttccc atatgtggct ctgtccaaga catacaatgt
     2041 agacaaacat gtgccagaca gtggagccac agccacggcc tacctgtgcg gggtcaaggg
     2101 caacttccag accattggct tgagtgcagc cgcccgcttt aaccagtgca acacgacacg
     2161 cggcaacgag gtcatctccg tgatgaatcg ggccaagaaa gcagggaagt cagtgggagt
     2221 ggtaaccacc acacgagtgc agcacgcctc gccagccggc acctacgccc acacggtgaa
     2281 ccgcaactgg tactcggacg ccgacgtgcc tgcctcggcc cgccaggagg ggtgccagga
     2341 catcgctacg cagctcatct ccaacatgga cattgacgtg atcctaggtg gaggccgaaa
     2401 gtacatgttt cgcatgggaa ccccagaccc tgagtaccca gatgactaca gccaaggtgg
     2461 gaccaggctg gacgggaaga atctggtgca ggaatggctg gcgaagcgcc agggtgcccg
     2521 gtatgtgtgg aaccgcactg agctcatgca ggcttccctg gacccgtctg tgacccatct
     2581 catgggtctc tttgagcctg gagacatgaa atacgagatc caccgagact ccacactgga
     2641 cccctccctg atggagatga cagaggctgc cctgcgcctg ctgagcagga acccccgcgg
     2701 cttcttcctc ttcgtggagg gtggtcgcat cgaccatggt catcatgaaa gcagggctta
     2761 ccgggcactg actgagacga tcatgttcga cgacgccatt gagagggcgg gccagctcac
     2821 cagcgaggag gacacgctga gcctcgtcac tgccgaccac tcccacgtct tctccttcgg
     2881 aggctacccc ctgcgaggga gctccatctt cgggctggcc cctggcaagg cccgggacag
     2941 gaaggcctac acggtcctcc tatacggaaa cggtccaggc tatgtgctca aggacggcgc
     3001 ccggccggat gttaccgaga gcgagagcgg gagccccgag tatcggcagc agtcagcagt
     3061 gcccctggac gaagagaccc acgcaggcga ggacgtggcg gtgttcgcgc gcggcccgca
     3121 ggcgcacctg gttcacggcg tgcaggagca gaccttcata gcgcacgtca tggccttcgc
     3181 cgcctgcctg gagccctaca ccgcctgcga cctggcgccc cccgccggca ccaccgacgc
     3241 cgcgcacccg ggtggcggag gactgaacga catcttcgag gctcagaaaa tcgaatggca
     3301 cgaacatcac catcaccatc accatcacca ttaaggccgg cctgtagtca ctggcacaac
     3361 gcgttacccg gtaagccaat cgggtataca cggtcgtcat actgcagaca gggttcttct
     3421 actttgcaag atagtcttga gtagtaaaat aaatagatag agaaaatcag cctcgactgt
     3481 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     3541 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcac aacactcaac
     3601 cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta
     3661 aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt
     3721 tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     3781 attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     3841 gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc
     3901 taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg
     3961 cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg
     4021 gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc ggatctgatc
     4081 agcacgtgtt gacaattaat catcggcata gtatatcggc atagtataat acgacaaggt
     4141 gaggaactaa accatgaccg agtacaagcc cacggtgcgc ctcgccaccc gcgacgacgt
     4201 ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac taccccgcca cgcgccacac
     4261 cgtcgatccg gaccgccaca tcgagcgggt caccgagctg caagaactct tcctcacgcg
     4321 cgtcgggctc gacatcggca aggtgtgggt cgcggacgac ggcgccgcgg tggcggtctg
     4381 gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc cgcgcatggc
     4441 cgagttgagc ggttcccggc tggccgcgca gcaacagatg gaaggcctcc tggcgccgca
     4501 ccggcccaag gagcccgcgt ggttcctggc caccgtcggc gtctcgcccg accaccaggg
     4561 caagggtctg ggcagcgccg tcgtgctccc cggagtggag gcggccgagc gcgccggggt
     4621 gcccgccttc ctggagacat ccgcgccccg caacctcccc ttctacgagc ggctcggctt
     4681 caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc acctggtgca tgacccgcaa
     4741 gcccggtgcc tgacacgtgc tacgagattt cgattccacc gccgccttct atgaaaggtt
     4801 gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat
     4861 gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag
     4921 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
     4981 gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt
     5041 ggcgtaatca tggtcattac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     5101 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     5161 gcttaccatc tggccccagc gctgcgatga taccgcgaga accacgctca ccggctccgg
     5221 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     5281 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     5341 ttaatagttt gcgcaacgtt gttgccatcg ctacaggcat cgtggtgtca cgctcgtcgt
     5401 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     5461 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     5521 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     5581 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     5641 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     5701 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     5761 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     5821 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     5881 agggaataag ggcgacacgg aaatgttgaa tactcatatt cttccttttt caatattatt
     5941 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     6001 ataaacaaat aggggtcagt gttacaacca attaaccaat tctgaacatt atcgcgagcc
     6061 catttatacc tgaatatggc tcataacacc ccttgctcat gaccaaaatc ccttaacgtg
     6121 agttacgcgc gcgtcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     6181 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     6241 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     6301 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag cccaccactt
     6361 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     6421 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     6481 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     6541 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     6601 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     6661 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     6721 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     6781 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     6841 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     6901 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag gcgagagtag
     6961 ggaactgcca ggcatcaaac taagcagaag gcccctgacg gatggccttt ttgcgtttct
     7021 acaaactctt tctgtgttgt aaaacgacgg ccagtcttaa gctcgggccc cctgggcggt
     7081 tctgataacg agtaatcgtt aatccgcaaa taacgtaaaa acccgcttcg gcgggttttt
     7141 ttatgggggg agtttaggga aagagcattt gtcagaatat ttaagggcgc ctgtcacttt
     7201 gcttgatata tgagaattat ttaaccttat aaatgagaaa aaagcaacgc actttaaata
     7261 agatacgttg ctttttcgat tgatgaacac ctataattaa actattcatc tattatttat
     7321 gattttttgt atatacaata tttctagttt gttaaagaga attaagaaaa taaatctcga
     7381 aaataataaa gggaaaatca gtttttgata tcaaaattat acatgtcaac gataatacaa
     7441 aatataatac aaactataag atgttatcag tatttattat catttagaat aaattttgtg
     7501 tcgcccttaa ttgtgagcgg ataacaatta cgagcttcat gcacagtggc gttgactcaa
     7561 tattggccat tagccatatt attcattggt tatatagcat aaatcaatat tggctattgg
     7621 ccattgcata cgttgtatct atatcataat atgtacattt atattggctc atgtccaata
     7681 tgaccgccat gtt
//
