LOCUS       sequence-417862-        8809 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 8809)
  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 8809)
  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-MDGA2-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..8809
                     /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"
     protein_bind    1889..1903
                     /label=boxB
                     /bound_moiety="lambda-N peptide"
                     /note="lambda N peptide binding site consisting of an RNA 
                     hairpin within the N-utliization (nut) site of the lambda 
                     bacteriophage transcript (Tan and Frankel, 1995)"
     CDS             4385..4429
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             4430..4447
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4587..4604)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4593..4704
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4822..4842)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4827..5156
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5007..5142
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        5204..5251
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5270..5869
                     /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(5270..5289)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5766..5786
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5999..6120
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6036..6055)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6090..6109
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(6173..6964,6965..7033))
                     /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     6796..6815
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7034..7125)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7167..7186
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     7192..7211
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      7297..7885
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7786..7805
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8039..8056
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     8154..8171
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     8155..8171
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    8627..8643
                     /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 ccacagttag aatcgtgcat tctggcctgg cctgcaacat
     1021 cgaggaagaa cggtacagcg agagagtgta caccatcaga gagggcgaga cactggaact
     1081 gacctgcctg gttaccggcc atcctcggcc tcagatcaga tggacaaaga cagccggcag
     1141 cgccagcgac agattccagg atagcagcgt gttcaacgaa accctgcgga tcaccaacat
     1201 ccagcggcat caaggcggac ggtactactg caaggccgag aatggactgg gcagccctgc
     1261 catcaagagc atcagagtgg acgtgtacta cctggacgac cccgtggtta cagtgcacca
     1321 gtctatcggc gaggccaaag agcagttcta ctacgagcgg accgtgttcc tgagatgcgt
     1381 ggccaacagc aatcctcctg tgcggtactc ttggcggaga ggccaagaag ttctgctgca
     1441 gggctctgac aaaggcgtgg aaatctacga gccattcttc acccaaggcg agacaaagat
     1501 cctgaagctg aagaacctgc ggcctcagga ctacgccaac tactcctgta ttgccagcgt
     1561 gcggaacgtg tgcaacatcc ccgataagat ggtgtccttc cggctgagca acaagaccgc
     1621 ctctcctagc atcaagctgc tggtggacga tcccatcgtg gtcaatcctg gcgaggctat
     1681 caccctcgtg tgtgtgacaa caggcggaga gcctgctcct agcctgacat gggtccgaag
     1741 ctttggcacc ctgcctgaga aaaccgtgct gaatggcggc accctgacaa tcccagccat
     1801 cacctctgat gacgccggca cctacagctg cattgccaac aacaacgtgg gcaaccccgc
     1861 caagaaaagc accaatatca tcgtgcgggc cctgaagaag ggcagattct ggatcacccc
     1921 tgatccttac cacaaggacg acaatatcca gatcggccgg gaagtgaaga tcagctgcca
     1981 ggtggaagcc gtgcctagcg aggaactgac attcagctgg ttcaagaacg gcagacccct
     2041 gagaagcagc gagcggatgg tcatcaccca gaccgatcct gatgtgtccc ctggcaccac
     2101 caacctggac atcatcgacc tgaagttcac cgacttcggc acctatacct gtgtggccag
     2161 cctgaaaggc ggcggaatca gcgacatcag catcgacgtg aacatcagca gcagcaccgt
     2221 gcctccaaac ctgaccgtgc cgcaagagaa gtctcctctg gtcacaagag aaggcgacac
     2281 catcgagctg cagtgtcaag tgaccggcaa gcccaagcct atcatcctgt ggtccagagc
     2341 cgacaaagag gtggccatgc cagatggcag catgcagatg gaaagctacg acggcacact
     2401 gcggatcgtg aacgtgtcca gagaaatgag cggcatgtac cggtgccaga ccagccagta
     2461 caacggcttt aacgtgaagc cccgggaagc cctggtgcag ctgattgtgc aatatcctcc
     2521 tgccgtggaa cccgccttcc tggaaatcag acagggccaa gaccgcagcg tgaccatgag
     2581 ctgtagagtg ctgcgggcct atcctatccg ggtgctgaca tatgagtggc gcctgggcaa
     2641 caaactgctg agaaccggcc agttcgactc tcaagagtac accgagtacg ccgtgaagtc
     2701 cctgagcaac gagaactacg gcgtgtacaa ctgcagcatc atcaacgagg ctggcgccgg
     2761 aagatgcagc tttctggtca ccggaaaggc ctacgctccc gagttttact acgacaccta
     2821 caatcccgtg tggcagaacc ggcaccgggt gtacagttac agcctgcagt ggacccagat
     2881 gaaccccgac gccgtggata ggatcgtggc ctatagactg ggcattagac aggccggaca
     2941 gcagcgttgg tgggagcaag agatcaagat caacgggaac atccagaagg gcgagctgat
     3001 cacctacaac ctgacagagc tgattaagcc cgaggcctac gaagtgcggc tgacccctct
     3061 gaccaagttt ggcgagggcg acagcaccat tagagtgatc aagtacagcg cccctgtgaa
     3121 ccctcacctg agagagttcc actgcggctt cgaggacggc aacatctgcc tgttcaccca
     3181 ggacgacacc gacaacttcg actggaccaa gcagagcacc gccaccagaa acaccaagta
     3241 cacccctaac acaggcccca acgccgacag aagcggcagc aaagagggct tctacatgta
     3301 catcgagaca agcagacccc ggctggaagg cgagaaagcc agactgctga gccccgtgtt
     3361 ttctatcgcc cctaagaacc cttacggccc caccaatacc gcctactgct tctccttctt
     3421 ctaccatatg tacggccagc acatcggagt gctgaacgtg tacctgagac tgaagggcca
     3481 gacaaccatc gagaaccctc tgtggtctag cagcggcaac aaaggccaga gatggaacga
     3541 ggcccatgtg aacatctacc ccatcaccag cttccagctg attttcgagg gcatcagagg
     3601 ccctggcatc gagggcgata ttgccatcga cgacgtgtcc attgccgagg gcgagtgtgc
     3661 caaacaggat ctggccacca agaatggcgc gccgggcggc gacaagaccc acacctgccc
     3721 cccctgccca gctccagaac tgctgggagg gccttccgtg ttcctgttcc cccccaagcc
     3781 caaggacacc ctgatgatca gcaggacccc cgaggtgacc tgcgtggtgg tggccgtgtc
     3841 ccacgaggac ccagaggtga agttcaactg gtacgtggac ggcgtggagg tgcacaacgc
     3901 caagaccaag cccagagagg agcagtacaa cagcacctac agggtggtgt ccgtgctgac
     3961 cgtgctgcac caggactggc tgaacggcaa agaatacaag tgcaaagtct ccaacaaggc
     4021 cctggctgcc ccaatcgaaa agacaatcag caaggccaag ggccagccac gggagcccca
     4081 ggtgtacacc ctgcccccca gccgggagga gatgaccaag aaccaggtgt ccctgacctg
     4141 tctggtgaag ggcttctacc ccagcgatat cgccgtggag tgggagagca acggccagcc
     4201 cgagaacaac tacaagacca cccccccagt gctggacagc gacggcagct tcttcctgta
     4261 cagcaagctg accgtggaca agtccaggtg gcagcagggc aacgtgttca gctgcagcgt
     4321 gatgcacgag gccctgcaca accactacac ccagaagtcc ctgagcctga gccccggcgg
     4381 cagcggtctt aatgatattt tcgaggctca gaagattgag tggcatgaac accaccacca
     4441 tcaccactaa ggccggcctg tagtcactgg cacaacgcgt tacccggtaa gccaatcggg
     4501 tatacacggt cgtcatactg cagacagggt tcttctactt tgcaagatag tcttgagtag
     4561 taaaataaat agatagagaa aatcagcctc gactgtgcct tctagttgcc agccatctgt
     4621 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     4681 ctaataaaat gaggaaattg catcacaaca ctcaacccta tctcggtcta ttcttttgat
     4741 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     4801 tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
     4861 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
     4921 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     4981 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     5041 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct
     5101 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
     5161 tcccgggagc ttgtatatcc attttcggat ctgatcagca cgtgttgaca attaatcatc
     5221 ggcatagtat atcggcatag tataatacga caaggtgagg aactaaacca tgaccgagta
     5281 caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac gcaccctcgc
     5341 cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc gatccggacc gccacatcga
     5401 gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt
     5461 gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc
     5521 gggggcggtg ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc
     5581 cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt
     5641 cctggccacc gtcggcgtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt
     5701 gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg agacatccgc
     5761 gccccgcaac ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt
     5821 gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgac acgtgctacg
     5881 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga
     5941 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa
     6001 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     6061 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     6121 tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt cattaccaat
     6181 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     6241 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagcgctg
     6301 cgatgatacc gcgagaacca cgctcaccgg ctccggattt atcagcaata aaccagccag
     6361 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     6421 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     6481 ccatcgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     6541 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     6601 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     6661 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     6721 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     6781 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     6841 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     6901 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     6961 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     7021 gttgaatact catattcttc ctttttcaat attattgaag catttatcag ggttattgtc
     7081 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gtcagtgtta
     7141 caaccaatta accaattctg aacattatcg cgagcccatt tatacctgaa tatggctcat
     7201 aacacccctt gctcatgacc aaaatccctt aacgtgagtt acgcgcgcgt cgttccactg
     7261 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     7321 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     7381 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     7441 tgttcttcta gtgtagccgt agttagccca ccacttcaag aactctgtag caccgcctac
     7501 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     7561 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     7621 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     7681 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     7741 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     7801 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     7861 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     7921 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     7981 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     8041 cgagtcagtg agcgaggaag cggaaggcga gagtagggaa ctgccaggca tcaaactaag
     8101 cagaaggccc ctgacggatg gcctttttgc gtttctacaa actctttctg tgttgtaaaa
     8161 cgacggccag tcttaagctc gggccccctg ggcggttctg ataacgagta atcgttaatc
     8221 cgcaaataac gtaaaaaccc gcttcggcgg gtttttttat ggggggagtt tagggaaaga
     8281 gcatttgtca gaatatttaa gggcgcctgt cactttgctt gatatatgag aattatttaa
     8341 ccttataaat gagaaaaaag caacgcactt taaataagat acgttgcttt ttcgattgat
     8401 gaacacctat aattaaacta ttcatctatt atttatgatt ttttgtatat acaatatttc
     8461 tagtttgtta aagagaatta agaaaataaa tctcgaaaat aataaaggga aaatcagttt
     8521 ttgatatcaa aattatacat gtcaacgata atacaaaata taatacaaac tataagatgt
     8581 tatcagtatt tattatcatt tagaataaat tttgtgtcgc ccttaattgt gagcggataa
     8641 caattacgag cttcatgcac agtggcgttg actcaatatt ggccattagc catattattc
     8701 attggttata tagcataaat caatattggc tattggccat tgcatacgtt gtatctatat
     8761 cataatatgt acatttatat tggctcatgt ccaatatgac cgccatgtt
//
