LOCUS       sequence-420965-        7879 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 7879)
  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 7879)
  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-EPCAM-COMP5AP-AviTag-9xHis
FEATURES             Location/Qualifiers
     source          1..7879
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    136..152
                     /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        319..698
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        699..902
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     853..873
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     899..923
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        1133..1151
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1133..1150
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     1164..1183
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             2036..2071
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             2081..2104
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             3764..3808
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             3809..3835
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     primer_bind     complement(3975..3992)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3981..4092
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4210..4230)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4215..4544
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4395..4530
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4457..4476
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4592..4639
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4658..5257
                     /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(4658..4677)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5154..5174
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5387..5508
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5424..5443)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5478..5497
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(5561..6352,6353..6421))
                     /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     6184..6203
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6422..6513)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6555..6574
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6580..6599
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      6685..7273
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7174..7193
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7427..7444
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     7542..7559
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7543..7559
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 ctcgaaaata ataaagggaa aatcagtttt tgatatcaaa attatacatg tcaacgataa
       61 tacaaaatat aatacaaact ataagatgtt atcagtattt attatcattt agaataaatt
      121 ttgtgtcgcc cttaattgtg agcggataac aattacgagc ttcatgcaca gtggcgttga
      181 ctcaatattg gccattagcc atattattca ttggttatat agcataaatc aatattggct
      241 attggccatt gcatacgttg tatctatatc ataatatgta catttatatt ggctcatgtc
      301 caatatgacc gccatgttgg cattgattat tgactagtta ttaatagtaa tcaattacgg
      361 ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc
      421 cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca
      481 tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg
      541 cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg
      601 acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt
      661 ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca
      721 tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg
      781 tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact
      841 ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag
      901 ctcgtttagt gaaccgtcag atcgcctgga gacgccatcc acgctgtttt gacctccata
      961 gaagacaccg ggaccgatcc agcctccgcg gccgggaacg gtgcattgga acgcggattc
     1021 cccgtgccaa gagtgacgta agtaccgcct atagagtcta taggcccacc cccttggctt
     1081 cgttagaacg cggctacaat taatacataa ccttatgtat catacacata cgatttaggt
     1141 gacactatag aataacatcc actttgcctt tctctccaca ggtgtccact cccaggtcca
     1201 actgcacctc ggttctatcg aaaacgcgcc tgctagcgcc accatgaaga ccatcattgc
     1261 cctgagctac atcttctgcc tggtgttcgc tgcggccgcc caagaggaat gcgtgtgcga
     1321 gaattacaag ctggccgtga actgcttcgt gaacaacaac agacagtgcc agtgtaccag
     1381 cgtgggagcc cagaataccg tgatctgtag caaactggcc gccaagtgcc tggtcatgaa
     1441 ggccgagatg aatggcagca agctcggcag aagggccaaa cctgaaggcg ccctgcagaa
     1501 caacgatggc ctgtacgacc ccgattgtga cgagtctggc ctgttcaagg ccaagcagtg
     1561 caacggcacc agcatgtgtt ggtgcgtgaa cacagccggc gtgcggagaa ccgacaagga
     1621 taccgagatc acctgtagcg agagagtgcg gacctactgg atcatcatcg agctgaagca
     1681 caaggccaga gagaagccct acgacagcaa gagcctgcgg acagctctgc agaaagagat
     1741 caccaccaga taccagctgg accccaagtt catcaccagc atcctgtacg agaacaacgt
     1801 gatcaccatc gacctggtgc agaacagcag ccagaaaacc cagaacgacg tggacattgc
     1861 cgacgtggcc tactacttcg agaaggacgt gaagggcgag agcctgttcc acagcaagaa
     1921 aatggacctg accgtgaacg gcgagcagct ggatcttgat cctggccaga cactgatcta
     1981 ctacgtggac gagaaggccc ctgagttcag catgcagggc ctgaaaggcg cgccggaaga
     2041 tcaagttgat ccaagattga ttgatggcaa gggcggatct ctggaggtgc tgttccaggg
     2101 accaggcgga agcggcaatc ccggcggcga tctggccccc cagatgctgc gcgagctgca
     2161 ggagaccaat gccgccctgc aggacgtgcg cgagctgctg cgccaccgcg tgaaggagat
     2221 caccttcctg aagaacaccg tgatggagtg cgacgcctgc ggtatgcagc cctctagagg
     2281 ctccggctcc atcatcccag ttgaggagga gaacccggac ttctggaacc gcgaggcagc
     2341 cgaggccctg ggtgccgcca agaagctgca gcctgcacag acagccgcca agaacctcat
     2401 catcttcctg ggcgatggga tgggggtgtc tacggtgaca gctgccagga tcctaaaagg
     2461 gcagaagaag gacaaactgg ggcctgagat acccctggcc atggaccgct tcccatatgt
     2521 ggctctgtcc aagacataca atgtagacaa acatgtgcca gacagtggag ccacagccac
     2581 ggcctacctg tgcggggtca agggcaactt ccagaccatt ggcttgagtg cagccgcccg
     2641 ctttaaccag tgcaacacga cacgcggcaa cgaggtcatc tccgtgatga atcgggccaa
     2701 gaaagcaggg aagtcagtgg gagtggtaac caccacacga gtgcagcacg cctcgccagc
     2761 cggcacctac gcccacacgg tgaaccgcaa ctggtactcg gacgccgacg tgcctgcctc
     2821 ggcccgccag gaggggtgcc aggacatcgc tacgcagctc atctccaaca tggacattga
     2881 cgtgatccta ggtggaggcc gaaagtacat gtttcgcatg ggaaccccag accctgagta
     2941 cccagatgac tacagccaag gtgggaccag gctggacggg aagaatctgg tgcaggaatg
     3001 gctggcgaag cgccagggtg cccggtatgt gtggaaccgc actgagctca tgcaggcttc
     3061 cctggacccg tctgtgaccc atctcatggg tctctttgag cctggagaca tgaaatacga
     3121 gatccaccga gactccacac tggacccctc cctgatggag atgacagagg ctgccctgcg
     3181 cctgctgagc aggaaccccc gcggcttctt cctcttcgtg gagggtggtc gcatcgacca
     3241 tggtcatcat gaaagcaggg cttaccgggc actgactgag acgatcatgt tcgacgacgc
     3301 cattgagagg gcgggccagc tcaccagcga ggaggacacg ctgagcctcg tcactgccga
     3361 ccactcccac gtcttctcct tcggaggcta ccccctgcga gggagctcca tcttcgggct
     3421 ggcccctggc aaggcccggg acaggaaggc ctacacggtc ctcctatacg gaaacggtcc
     3481 aggctatgtg ctcaaggacg gcgcccggcc ggatgttacc gagagcgaga gcgggagccc
     3541 cgagtatcgg cagcagtcag cagtgcccct ggacgaagag acccacgcag gcgaggacgt
     3601 ggcggtgttc gcgcgcggcc cgcaggcgca cctggttcac ggcgtgcagg agcagacctt
     3661 catagcgcac gtcatggcct tcgccgcctg cctggagccc tacaccgcct gcgacctggc
     3721 gccccccgcc ggcaccaccg acgccgcgca cccgggtggc ggaggactga acgacatctt
     3781 cgaggctcag aaaatcgaat ggcacgaaca tcaccatcac catcaccatc accattaagg
     3841 ccggcctgta gtcactggca caacgcgtta cccggtaagc caatcgggta tacacggtcg
     3901 tcatactgca gacagggttc ttctactttg caagatagtc ttgagtagta aaataaatag
     3961 atagagaaaa tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
     4021 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     4081 ggaaattgca tcacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     4141 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     4201 taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     4261 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     4321 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     4381 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     4441 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc
     4501 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt
     4561 gtatatccat tttcggatct gatcagcacg tgttgacaat taatcatcgg catagtatat
     4621 cggcatagta taatacgaca aggtgaggaa ctaaaccatg accgagtaca agcccacggt
     4681 gcgcctcgcc acccgcgacg acgtccccag ggccgtacgc accctcgccg ccgcgttcgc
     4741 cgactacccc gccacgcgcc acaccgtcga tccggaccgc cacatcgagc gggtcaccga
     4801 gctgcaagaa ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga
     4861 cgacggcgcc gcggtggcgg tctggaccac gccggagagc gtcgaagcgg gggcggtgtt
     4921 cgccgagatc ggcccgcgca tggccgagtt gagcggttcc cggctggccg cgcagcaaca
     4981 gatggaaggc ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc tggccaccgt
     5041 cggcgtctcg cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc tccccggagt
     5101 ggaggcggcc gagcgcgccg gggtgcccgc cttcctggag acatccgcgc cccgcaacct
     5161 ccccttctac gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc
     5221 gcgcacctgg tgcatgaccc gcaagcccgg tgcctgacac gtgctacgag atttcgattc
     5281 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     5341 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc
     5401 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     5461 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat
     5521 accgtcgacc tctagctaga gcttggcgta atcatggtca ttaccaatgc ttaatcagtg
     5581 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     5641 tgtagataac tacgatacgg gagggcttac catctggccc cagcgctgcg atgataccgc
     5701 gagaaccacg ctcaccggct ccggatttat cagcaataaa ccagccagcc ggaagggccg
     5761 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     5821 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc atcgctacag
     5881 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     5941 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     6001 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     6061 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     6121 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     6181 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     6241 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     6301 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     6361 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     6421 tattcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     6481 acatatttga atgtatttag aaaaataaac aaataggggt cagtgttaca accaattaac
     6541 caattctgaa cattatcgcg agcccattta tacctgaata tggctcataa caccccttgc
     6601 tcatgaccaa aatcccttaa cgtgagttac gcgcgcgtcg ttccactgag cgtcagaccc
     6661 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     6721 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     6781 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     6841 gtagccgtag ttagcccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     6901 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     6961 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     7021 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     7081 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     7141 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     7201 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     7261 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     7321 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     7381 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     7441 cgaggaagcg gaaggcgaga gtagggaact gccaggcatc aaactaagca gaaggcccct
     7501 gacggatggc ctttttgcgt ttctacaaac tctttctgtg ttgtaaaacg acggccagtc
     7561 ttaagctcgg gccccctggg cggttctgat aacgagtaat cgttaatccg caaataacgt
     7621 aaaaacccgc ttcggcgggt ttttttatgg ggggagttta gggaaagagc atttgtcaga
     7681 atatttaagg gcgcctgtca ctttgcttga tatatgagaa ttatttaacc ttataaatga
     7741 gaaaaaagca acgcacttta aataagatac gttgcttttt cgattgatga acacctataa
     7801 ttaaactatt catctattat ttatgatttt ttgtatatac aatatttcta gtttgttaaa
     7861 gagaattaag aaaataaat
//
