LOCUS       sequence-417861-        6832 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of cell-surface protein extracellular domain 
            fused to Fc(DAPA)-Avi-6xHis. Protein is secreted from cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6832)
  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 6832)
  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-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6832
                     /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             2408..2452
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             2453..2470
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2610..2627)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2616..2727
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2845..2865)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2850..3179
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3030..3165
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3092..3111
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3227..3274
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3293..3892
                     /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(3293..3312)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3789..3809
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4022..4143
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4059..4078)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4113..4132
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(4196..4987,4988..5056))
                     /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     4819..4838
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5057..5148)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5190..5209
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5215..5234
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      5320..5908
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5809..5828
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6062..6079
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     6177..6194
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6178..6194
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6650..6666
                     /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 cccaagagga atgcgtgtgc gagaattaca agctggccgt
     1021 gaactgcttc gtgaacaaca acagacagtg ccagtgtacc agcgtgggag cccagaatac
     1081 cgtgatctgt agcaaactgg ccgccaagtg cctggtcatg aaggccgaga tgaatggcag
     1141 caagctcggc agaagggcca aacctgaagg cgccctgcag aacaacgatg gcctgtacga
     1201 ccccgattgt gacgagtctg gcctgttcaa ggccaagcag tgcaacggca ccagcatgtg
     1261 ttggtgcgtg aacacagccg gcgtgcggag aaccgacaag gataccgaga tcacctgtag
     1321 cgagagagtg cggacctact ggatcatcat cgagctgaag cacaaggcca gagagaagcc
     1381 ctacgacagc aagagcctgc ggacagctct gcagaaagag atcaccacca gataccagct
     1441 ggaccccaag ttcatcacca gcatcctgta cgagaacaac gtgatcacca tcgacctggt
     1501 gcagaacagc agccagaaaa cccagaacga cgtggacatt gccgacgtgg cctactactt
     1561 cgagaaggac gtgaagggcg agagcctgtt ccacagcaag aaaatggacc tgaccgtgaa
     1621 cggcgagcag ctggatcttg atcctggcca gacactgatc tactacgtgg acgagaaggc
     1681 ccctgagttc agcatgcagg gcctgaaagg cgcgccgggc ggcgacaaga cccacacctg
     1741 ccccccctgc ccagctccag aactgctggg agggccttcc gtgttcctgt tcccccccaa
     1801 gcccaaggac accctgatga tcagcaggac ccccgaggtg acctgcgtgg tggtggccgt
     1861 gtcccacgag gacccagagg tgaagttcaa ctggtacgtg gacggcgtgg aggtgcacaa
     1921 cgccaagacc aagcccagag aggagcagta caacagcacc tacagggtgg tgtccgtgct
     1981 gaccgtgctg caccaggact ggctgaacgg caaagaatac aagtgcaaag tctccaacaa
     2041 ggccctggct gccccaatcg aaaagacaat cagcaaggcc aagggccagc cacgggagcc
     2101 ccaggtgtac accctgcccc ccagccggga ggagatgacc aagaaccagg tgtccctgac
     2161 ctgtctggtg aagggcttct accccagcga tatcgccgtg gagtgggaga gcaacggcca
     2221 gcccgagaac aactacaaga ccaccccccc agtgctggac agcgacggca gcttcttcct
     2281 gtacagcaag ctgaccgtgg acaagtccag gtggcagcag ggcaacgtgt tcagctgcag
     2341 cgtgatgcac gaggccctgc acaaccacta cacccagaag tccctgagcc tgagccccgg
     2401 cggcagcggt cttaatgata ttttcgaggc tcagaagatt gagtggcatg aacaccacca
     2461 ccatcaccac taaggccggc ctgtagtcac tggcacaacg cgttacccgg taagccaatc
     2521 gggtatacac ggtcgtcata ctgcagacag ggttcttcta ctttgcaaga tagtcttgag
     2581 tagtaaaata aatagataga gaaaatcagc ctcgactgtg ccttctagtt gccagccatc
     2641 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
     2701 ttcctaataa aatgaggaaa ttgcatcaca acactcaacc ctatctcggt ctattctttt
     2761 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     2821 aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
     2881 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
     2941 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     3001 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     3061 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg
     3121 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
     3181 agctcccggg agcttgtata tccattttcg gatctgatca gcacgtgttg acaattaatc
     3241 atcggcatag tatatcggca tagtataata cgacaaggtg aggaactaaa ccatgaccga
     3301 gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct
     3361 cgccgccgcg ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat
     3421 cgagcgggtc accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa
     3481 ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga
     3541 agcgggggcg gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct
     3601 ggccgcgcag caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg
     3661 gttcctggcc accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt
     3721 cgtgctcccc ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacatc
     3781 cgcgccccgc aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga
     3841 ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag cccggtgcct gacacgtgct
     3901 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg
     3961 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc
     4021 caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     4081 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     4141 ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcattacc
     4201 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     4261 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagcg
     4321 ctgcgatgat accgcgagaa ccacgctcac cggctccgga tttatcagca ataaaccagc
     4381 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     4441 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     4501 ttgccatcgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     4561 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     4621 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     4681 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     4741 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     4801 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     4861 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     4921 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     4981 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     5041 aatgttgaat actcatattc ttcctttttc aatattattg aagcatttat cagggttatt
     5101 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggtcagtg
     5161 ttacaaccaa ttaaccaatt ctgaacatta tcgcgagccc atttatacct gaatatggct
     5221 cataacaccc cttgctcatg accaaaatcc cttaacgtga gttacgcgcg cgtcgttcca
     5281 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     5341 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     5401 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     5461 tactgttctt ctagtgtagc cgtagttagc ccaccacttc aagaactctg tagcaccgcc
     5521 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     5581 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     5641 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     5701 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     5761 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     5821 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     5881 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     5941 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     6001 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     6061 cagcgagtca gtgagcgagg aagcggaagg cgagagtagg gaactgccag gcatcaaact
     6121 aagcagaagg cccctgacgg atggcctttt tgcgtttcta caaactcttt ctgtgttgta
     6181 aaacgacggc cagtcttaag ctcgggcccc ctgggcggtt ctgataacga gtaatcgtta
     6241 atccgcaaat aacgtaaaaa cccgcttcgg cgggtttttt tatgggggga gtttagggaa
     6301 agagcatttg tcagaatatt taagggcgcc tgtcactttg cttgatatat gagaattatt
     6361 taaccttata aatgagaaaa aagcaacgca ctttaaataa gatacgttgc tttttcgatt
     6421 gatgaacacc tataattaaa ctattcatct attatttatg attttttgta tatacaatat
     6481 ttctagtttg ttaaagagaa ttaagaaaat aaatctcgaa aataataaag ggaaaatcag
     6541 tttttgatat caaaattata catgtcaacg ataatacaaa atataataca aactataaga
     6601 tgttatcagt atttattatc atttagaata aattttgtgt cgcccttaat tgtgagcgga
     6661 taacaattac gagcttcatg cacagtggcg ttgactcaat attggccatt agccatatta
     6721 ttcattggtt atatagcata aatcaatatt ggctattggc cattgcatac gttgtatcta
     6781 tatcataata tgtacattta tattggctca tgtccaatat gaccgccatg tt
//
