LOCUS       sequence-304363-        8866 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 8866)
  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 8866)
  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-mIslr2-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..8866
                     /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             4471..4506
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             4516..4539
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             6199..6243
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6244..6270
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(6410..6427)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6416..6527
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6645..6665)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6650..6979
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6830..6965
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6892..6911
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7027..7074
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7093..7692
                     /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(7093..7112)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7589..7609
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7822..7943
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7859..7878)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7913..7932
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7996..8787,8788..8856))
                     /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     8619..8638
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8857..8866,1..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 cggccgccgc gtgtcccgaa
     2761 ccttgtgctt gcgtagataa atacgcgcac cagttcgcgg actgtgctta caaggaactc
     2821 cgcgaggttc cagaaggcct tcctgccaac gtgaccacac tcagtctgtc tgctaacaaa
     2881 attactgtgc ttcgacgagg agcatttgta aatgtaactc aagtaacatc cctttggctt
     2941 gcacacagcg aagtgaggac ggtcgaaagt ggtgcacttg ccgtactttc acaattgaaa
     3001 aatctcgatc tgtcccacaa cttgatttcc aattttccgt ggagcgactt gcggaatttg
     3061 agcgcgcttc agctgctgaa gatgaatcac aatcggctgg ggtccctccc acgagacgcg
     3121 cttggcgccc tgcccgacct taggagcctt cggatcaata ataatcgcct ccggaccctg
     3181 gagcccggta catttgacgc actcagcgcg ctctctcacc ttcaacttta tcataatccc
     3241 ttccattgca gttgtggatt ggtgtggttg caagcctggg ctgcaagtac acgcgtcagc
     3301 ttgccagagc ccgacagtat cgcgtgcgcg agtccacctg agttgcaagg ggtaccggtc
     3361 cacaggttgc cagcgctgcc atgtgccccg ccaagcgtcc gactcagtgc tgagccacca
     3421 cctgaagcgc cggggacacc acttcgcgcc ggactcgcgt tcatgctgca ttgtgtggcc
     3481 gagggccatc ctacaccgag gttgcaatgg cagctgcaga tcccgggggg aactgtagtt
     3541 ctcgttcctc ccgttctctc caaggaggag gatgggggcg acaaggtcga agatggggaa
     3601 ggcgacggag atgaggacct tccgacccaa acggaggcgc caactccaac gccagcaccg
     3661 gcatggcccg ctccgcctgc cacgccgcgc tttttggccc ttgcaaacgg atccctgttg
     3721 gtccctctgc tctctgcaaa agaggcgggg atttacacct gtagggccca taatgagttg
     3781 ggtaccaatt ctaccagcct ccgagtgaca gtcgcagcag ccgggccccc caaacatgct
     3841 ccagggaccg gcgaggagcc agacgcacaa gtccctacat ctgaaaggaa agctacaaca
     3901 aagggtaggt ctaacagtgt acttccgttc aaacctgagg gtaagacaaa agggcagggg
     3961 ttggcgaggg ttagcgtact gggcgaaatt gaagcagagt tggaagaaac agatgaagga
     4021 gaacaaatgg aggggcagat cccagctgac cccatgggag aaaaacactg tgggcacgga
     4081 gatccatctc ggtatgtcag taatcatgct ttcaaccaat caagcgacct caaaccccac
     4141 gtttttgaac tcggcgtaat cgcacttgac gtggcagagc gggaagcgag agtacaactg
     4201 accccacttg ccgcccgatg gggaccaggg cccgatgggg cctcaggcgc gagaaggcca
     4261 ggcagacgac ctctccggct tctctacctg tgtccagcgg gtggtggcac tgcagtgcag
     4321 tggagtaggg tggaggaggg tgtgaatgct tattggttta ggggccttcg ccctggtact
     4381 aattactctg tctgtttggc gcttgccgga gaggcttgtc atgttcaggt agtatttagc
     4441 actaaaaaag aactcccgag tggcgcgccg gaagatcaag ttgatccaag attgattgat
     4501 ggcaagggcg gatctctgga ggtgctgttc cagggaccag gcggaagcgg caatcccggc
     4561 ggcgatctgg ccccccagat gctgcgcgag ctgcaggaga ccaatgccgc cctgcaggac
     4621 gtgcgcgagc tgctgcgcca ccgcgtgaag gagatcacct tcctgaagaa caccgtgatg
     4681 gagtgcgacg cctgcggtat gcagccctct agaggctccg gctccatcat cccagttgag
     4741 gaggagaacc cggacttctg gaaccgcgag gcagccgagg ccctgggtgc cgccaagaag
     4801 ctgcagcctg cacagacagc cgccaagaac ctcatcatct tcctgggcga tgggatgggg
     4861 gtgtctacgg tgacagctgc caggatccta aaagggcaga agaaggacaa actggggcct
     4921 gagatacccc tggccatgga ccgcttccca tatgtggctc tgtccaagac atacaatgta
     4981 gacaaacatg tgccagacag tggagccaca gccacggcct acctgtgcgg ggtcaagggc
     5041 aacttccaga ccattggctt gagtgcagcc gcccgcttta accagtgcaa cacgacacgc
     5101 ggcaacgagg tcatctccgt gatgaatcgg gccaagaaag cagggaagtc agtgggagtg
     5161 gtaaccacca cacgagtgca gcacgcctcg ccagccggca cctacgccca cacggtgaac
     5221 cgcaactggt actcggacgc cgacgtgcct gcctcggccc gccaggaggg gtgccaggac
     5281 atcgctacgc agctcatctc caacatggac attgacgtga tcctaggtgg aggccgaaag
     5341 tacatgtttc gcatgggaac cccagaccct gagtacccag atgactacag ccaaggtggg
     5401 accaggctgg acgggaagaa tctggtgcag gaatggctgg cgaagcgcca gggtgcccgg
     5461 tatgtgtgga accgcactga gctcatgcag gcttccctgg acccgtctgt gacccatctc
     5521 atgggtctct ttgagcctgg agacatgaaa tacgagatcc accgagactc cacactggac
     5581 ccctccctga tggagatgac agaggctgcc ctgcgcctgc tgagcaggaa cccccgcggc
     5641 ttcttcctct tcgtggaggg tggtcgcatc gaccatggtc atcatgaaag cagggcttac
     5701 cgggcactga ctgagacgat catgttcgac gacgccattg agagggcggg ccagctcacc
     5761 agcgaggagg acacgctgag cctcgtcact gccgaccact cccacgtctt ctccttcgga
     5821 ggctaccccc tgcgagggag ctccatcttc gggctggccc ctggcaaggc ccgggacagg
     5881 aaggcctaca cggtcctcct atacggaaac ggtccaggct atgtgctcaa ggacggcgcc
     5941 cggccggatg ttaccgagag cgagagcggg agccccgagt atcggcagca gtcagcagtg
     6001 cccctggacg aagagaccca cgcaggcgag gacgtggcgg tgttcgcgcg cggcccgcag
     6061 gcgcacctgg ttcacggcgt gcaggagcag accttcatag cgcacgtcat ggccttcgcc
     6121 gcctgcctgg agccctacac cgcctgcgac ctggcgcccc ccgccggcac caccgacgcc
     6181 gcgcacccgg gtggcggagg actgaacgac atcttcgagg ctcagaaaat cgaatggcac
     6241 gaacatcacc atcaccatca ccatcaccat taaggccggc ctgtagtcac tggcacaacg
     6301 cgttacccgg taagccaatc gggtatacac ggtcgtcata ctgcagacag ggttcttcta
     6361 ctttgcaaga tagtcttgag tagtaaaata aatagataga gaaaatcagc ctcgactgtg
     6421 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
     6481 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcaca acactcaacc
     6541 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     6601 aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt
     6661 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     6721 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     6781 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     6841 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     6901 agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg
     6961 aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca
     7021 gcacgtgttg acaattaatc atcggcatag tatatcggca tagtataata cgacaaggtg
     7081 aggaactaaa ccatgaccga gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc
     7141 cccagggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac gcgccacacc
     7201 gtcgatccgg accgccacat cgagcgggtc accgagctgc aagaactctt cctcacgcgc
     7261 gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg
     7321 accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc gcgcatggcc
     7381 gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct ggcgccgcac
     7441 cggcccaagg agcccgcgtg gttcctggcc accgtcggcg tctcgcccga ccaccagggc
     7501 aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg cgccggggtg
     7561 cccgccttcc tggagacatc cgcgccccgc aacctcccct tctacgagcg gctcggcttc
     7621 accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag
     7681 cccggtgcct gacacgtgct acgagatttc gattccaccg ccgccttcta tgaaaggttg
     7741 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     7801 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
     7861 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     7921 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
     7981 gcgtaatcat ggtcattacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     8041 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     8101 cttaccatct ggccccagcg ctgcgatgat accgcgagaa ccacgctcac cggctccgga
     8161 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     8221 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     8281 taatagtttg cgcaacgttg ttgccatcgc tacaggcatc gtggtgtcac gctcgtcgtt
     8341 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     8401 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     8461 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     8521 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     8581 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     8641 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     8701 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     8761 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     8821 gggaataagg gcgacacgga aatgttgaat actcatattc ttcctt
//
