LOCUS       sequence-449664-        8467 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 8467)
  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 8467)
  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-CD276-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8467
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     124..143
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     149..168
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      254..842
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     743..762
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     996..1013
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     1111..1128
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1112..1128
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1584..1600
                     /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        1767..2146
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2147..2350
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2301..2321
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2347..2371
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        2581..2599
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2581..2598
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     2612..2631
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             4072..4107
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             4117..4140
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             5800..5844
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5845..5871
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(6011..6028)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6017..6128
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6246..6266)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6251..6580
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6431..6566
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6493..6512
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6628..6675
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6694..7293
                     /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(6694..6713)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7190..7210
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7423..7544
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7460..7479)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7514..7533
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7597..8388,8389..8457))
                     /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     8220..8239
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8458..8467,1..82))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
       61 tgtatttaga aaaataaaca aataggggtc agtgttacaa ccaattaacc aattctgaac
      121 attatcgcga gcccatttat acctgaatat ggctcataac accccttgct catgaccaaa
      181 atcccttaac gtgagttacg cgcgcgtcgt tccactgagc gtcagacccc gtagaaaaga
      241 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
      301 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
      361 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
      421 tagcccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
      481 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
      541 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
      601 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
      661 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
      721 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
      781 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
      841 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
      901 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag
      961 ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg
     1021 aaggcgagag tagggaactg ccaggcatca aactaagcag aaggcccctg acggatggcc
     1081 tttttgcgtt tctacaaact ctttctgtgt tgtaaaacga cggccagtct taagctcggg
     1141 ccccctgggc ggttctgata acgagtaatc gttaatccgc aaataacgta aaaacccgct
     1201 tcggcgggtt tttttatggg gggagtttag ggaaagagca tttgtcagaa tatttaaggg
     1261 cgcctgtcac tttgcttgat atatgagaat tatttaacct tataaatgag aaaaaagcaa
     1321 cgcactttaa ataagatacg ttgctttttc gattgatgaa cacctataat taaactattc
     1381 atctattatt tatgattttt tgtatataca atatttctag tttgttaaag agaattaaga
     1441 aaataaatct cgaaaataat aaagggaaaa tcagtttttg atatcaaaat tatacatgtc
     1501 aacgataata caaaatataa tacaaactat aagatgttat cagtatttat tatcatttag
     1561 aataaatttt gtgtcgccct taattgtgag cggataacaa ttacgagctt catgcacagt
     1621 ggcgttgact caatattggc cattagccat attattcatt ggttatatag cataaatcaa
     1681 tattggctat tggccattgc atacgttgta tctatatcat aatatgtaca tttatattgg
     1741 ctcatgtcca atatgaccgc catgttggca ttgattattg actagttatt aatagtaatc
     1801 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
     1861 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
     1921 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
     1981 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
     2041 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
     2101 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
     2161 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
     2221 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
     2281 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
     2341 aagcagagct cgtttagtga accgtcagat cgcctggaga cgccatccac gctgttttga
     2401 cctccataga agacaccggg accgatccag cctccgcggc cgggaacggt gcattggaac
     2461 gcggattccc cgtgccaaga gtgacgtaag taccgcctat agagtctata ggcccacccc
     2521 cttggcttcg ttagaacgcg gctacaatta atacataacc ttatgtatca tacacatacg
     2581 atttaggtga cactatagaa taacatccac tttgcctttc tctccacagg tgtccactcc
     2641 caggtccaac tgcacctcgg ttctatcgaa aacgcgcctg ctagcgccac catgaagacc
     2701 atcattgccc tgagctacat cttctgcctg gtgttcgctg cggccgcctt ggaagtgcaa
     2761 gtgcctgaag atcctgtggt ggccctcgtg ggaacagatg ccacactgtg ctgtagcttt
     2821 agccccgagc ctggattttc tctggcccag ctgaatctga tctggcagct gaccgatacc
     2881 aagcagctgg tgcactcttt cgccgaaggc caggatcagg gcagcgccta tgctaacaga
     2941 accgctctgt ttcccgatct gctggcccag ggaaatgcca gcctgagact gcagagagtc
     3001 agagttgccg acgagggcag cttcacctgt ttcgtgtcca tcagagattt cggctctgcc
     3061 gccgtgtctc tgcaagtggc cgctccttac agcaagccca gcatgaccct ggaacctaac
     3121 aaggacctga ggcctggcga caccgtgacc atcacatgta gcagctacca gggctatccc
     3181 gaggccgaag tgttttggca ggatggacaa ggcgtgcccc tgaccggcaa tgtgacaaca
     3241 tctcagatgg ccaacgagca gggcctgttc gatgtgcact ctatcctgag agtggtgctg
     3301 ggcgccaatg gcacatatag ctgtctcgtg cggaaccccg ttctgcagca ggatgcccac
     3361 agcagcgtga caatcacacc ccagagatct ccaacaggcg ccgtggaagt tcaggtgcca
     3421 gaggaccctg ttgtggctct tgtgggcacc gatgctaccc tgagatgctc ttttagccct
     3481 gaaccaggct tcagcctcgc tcagctgaac ctcatttggc aactcacaga cacaaaacag
     3541 ctcgtccact ccttcaccga gggaagagat cagggctccg cctacgccaa taggacagca
     3601 ctgtttccag acctgctcgc tcaaggcaac gctagtctga ggctgcaacg cgttagagtg
     3661 gccgatgaag gctcctttac ctgcttcgtc agcatccgcg attttggcag tgctgccgtc
     3721 agcctgcaag tcgctgcccc ttactccaag ccttctatga ccctcgagcc gaacaaagat
     3781 ctcagacccg gcgatacagt gaccattacc tgcagctcct acagaggcta cccagaggct
     3841 gaggttttct ggcaagacgg acagggcgtt ccactgactg gaaacgtgac caccagccag
     3901 atggctaatg aacagggact ctttgacgtg cacagcgtgc tgagagtcgt tctgggagct
     3961 aacggcacct attcttgcct cgtcagaaac cctgtgctcc agcaggacgc tcacggctct
     4021 gtgacaatta ccggccagcc tatgaccttt ccacctgagg ctggcgcgcc ggaagatcaa
     4081 gttgatccaa gattgattga tggcaagggc ggatctctgg aggtgctgtt ccagggacca
     4141 ggcggaagcg gcaatcccgg cggcgatctg gccccccaga tgctgcgcga gctgcaggag
     4201 accaatgccg ccctgcagga cgtgcgcgag ctgctgcgcc accgcgtgaa ggagatcacc
     4261 ttcctgaaga acaccgtgat ggagtgcgac gcctgcggta tgcagccctc tagaggctcc
     4321 ggctccatca tcccagttga ggaggagaac ccggacttct ggaaccgcga ggcagccgag
     4381 gccctgggtg ccgccaagaa gctgcagcct gcacagacag ccgccaagaa cctcatcatc
     4441 ttcctgggcg atgggatggg ggtgtctacg gtgacagctg ccaggatcct aaaagggcag
     4501 aagaaggaca aactggggcc tgagataccc ctggccatgg accgcttccc atatgtggct
     4561 ctgtccaaga catacaatgt agacaaacat gtgccagaca gtggagccac agccacggcc
     4621 tacctgtgcg gggtcaaggg caacttccag accattggct tgagtgcagc cgcccgcttt
     4681 aaccagtgca acacgacacg cggcaacgag gtcatctccg tgatgaatcg ggccaagaaa
     4741 gcagggaagt cagtgggagt ggtaaccacc acacgagtgc agcacgcctc gccagccggc
     4801 acctacgccc acacggtgaa ccgcaactgg tactcggacg ccgacgtgcc tgcctcggcc
     4861 cgccaggagg ggtgccagga catcgctacg cagctcatct ccaacatgga cattgacgtg
     4921 atcctaggtg gaggccgaaa gtacatgttt cgcatgggaa ccccagaccc tgagtaccca
     4981 gatgactaca gccaaggtgg gaccaggctg gacgggaaga atctggtgca ggaatggctg
     5041 gcgaagcgcc agggtgcccg gtatgtgtgg aaccgcactg agctcatgca ggcttccctg
     5101 gacccgtctg tgacccatct catgggtctc tttgagcctg gagacatgaa atacgagatc
     5161 caccgagact ccacactgga cccctccctg atggagatga cagaggctgc cctgcgcctg
     5221 ctgagcagga acccccgcgg cttcttcctc ttcgtggagg gtggtcgcat cgaccatggt
     5281 catcatgaaa gcagggctta ccgggcactg actgagacga tcatgttcga cgacgccatt
     5341 gagagggcgg gccagctcac cagcgaggag gacacgctga gcctcgtcac tgccgaccac
     5401 tcccacgtct tctccttcgg aggctacccc ctgcgaggga gctccatctt cgggctggcc
     5461 cctggcaagg cccgggacag gaaggcctac acggtcctcc tatacggaaa cggtccaggc
     5521 tatgtgctca aggacggcgc ccggccggat gttaccgaga gcgagagcgg gagccccgag
     5581 tatcggcagc agtcagcagt gcccctggac gaagagaccc acgcaggcga ggacgtggcg
     5641 gtgttcgcgc gcggcccgca ggcgcacctg gttcacggcg tgcaggagca gaccttcata
     5701 gcgcacgtca tggccttcgc cgcctgcctg gagccctaca ccgcctgcga cctggcgccc
     5761 cccgccggca ccaccgacgc cgcgcacccg ggtggcggag gactgaacga catcttcgag
     5821 gctcagaaaa tcgaatggca cgaacatcac catcaccatc accatcacca ttaaggccgg
     5881 cctgtagtca ctggcacaac gcgttacccg gtaagccaat cgggtataca cggtcgtcat
     5941 actgcagaca gggttcttct actttgcaag atagtcttga gtagtaaaat aaatagatag
     6001 agaaaatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc
     6061 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa
     6121 attgcatcac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     6181 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat
     6241 tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag
     6301 tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc
     6361 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct
     6421 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg
     6481 actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa
     6541 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat
     6601 atccattttc ggatctgatc agcacgtgtt gacaattaat catcggcata gtatatcggc
     6661 atagtataat acgacaaggt gaggaactaa accatgaccg agtacaagcc cacggtgcgc
     6721 ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac
     6781 taccccgcca cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg
     6841 caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac
     6901 ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc
     6961 gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg
     7021 gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcggc
     7081 gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag
     7141 gcggccgagc gcgccggggt gcccgccttc ctggagacat ccgcgccccg caacctcccc
     7201 ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc
     7261 acctggtgca tgacccgcaa gcccggtgcc tgacacgtgc tacgagattt cgattccacc
     7321 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc
     7381 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct
     7441 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     7501 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg
     7561 tcgacctcta gctagagctt ggcgtaatca tggtcattac caatgcttaa tcagtgaggc
     7621 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     7681 gataactacg atacgggagg gcttaccatc tggccccagc gctgcgatga taccgcgaga
     7741 accacgctca ccggctccgg atttatcagc aataaaccag ccagccggaa gggccgagcg
     7801 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     7861 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccatcg ctacaggcat
     7921 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     7981 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     8041 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     8101 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     8161 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     8221 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     8281 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     8341 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     8401 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatatt
     8461 cttcctt
//
