LOCUS       sequence-368521-        6571 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 6571)
  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 6571)
  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-TNFRSF10B-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6571
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     169..189
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    307..418
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     407..424
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(564..581)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(582..626)
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     primer_bind     complement(1908..1927)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        complement(1940..1958)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1941..1958)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2168..2192)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        complement(2189..2392)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2218..2238)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2393..2772
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     protein_bind    2939..2955
                     /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)."
     primer_bind     complement(3411..3428)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3411..3427)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3526..3543)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3697..4285)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3777..3796)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4371..4390)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(4396..4415)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        4457..4548
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4549..4617,4618..5409)
                     /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     complement(4767..4786)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     polyA_signal    5462..5583
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5473..5492)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5527..5546
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(5713..6312)
                     /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(5796..5816)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     6293..6312
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(6331..6378)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(join(6426..6571,1..184))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(6440..6571,1..4)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(6494..6513)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
ORIGIN
        1 ggatgggcgg agttaggggc gggactatgg ttgctgacta attgagatgc atgctttgca
       61 tacttctgcc tgctggggag cctggggact ttccacacct ggttgctgac taattgagat
      121 gcatgctttg catacttctg cctgctgggg agcctgggga ctttccacac cctaactgac
      181 acacattcca cagaattaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca
      241 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag
      301 tgttgtgatg caatttcctc attttattag gaaaggacag tgggagtggc accttccagg
      361 gtcaaggaag gcacggggga ggggcaaaca acagatggct ggcaactaga aggcacagtc
      421 gaggctgatt ttctctatct atttatttta ctactcaaga ctatcttgca aagtagaaga
      481 accctgtctg cagtatgacg accgtgtata cccgattggc ttaccgggta acgcgttgtg
      541 ccagtgacta caggccggcc ttagtggtga tggtggtggt gttcatgcca ctcaatcttc
      601 tgagcctcga aaatatcatt aagaccgctg ccgccggggc tcaggctcag ggacttctgg
      661 gtgtagtggt tgtgcagggc ctcgtgcatc acgctgcagc tgaacacgtt gccctgctgc
      721 cacctggact tgtccacggt cagcttgctg tacaggaaga agctgccgtc gctgtccagc
      781 actggggggg tggtcttgta gttgttctcg ggctggccgt tgctctccca ctccacggcg
      841 atatcgctgg ggtagaagcc cttcaccaga caggtcaggg acacctggtt cttggtcatc
      901 tcctcccggc tggggggcag ggtgtacacc tggggctccc gtggctggcc cttggccttg
      961 ctgattgtct tttcgattgg ggcagccagg gccttgttgg agactttgca cttgtattct
     1021 ttgccgttca gccagtcctg gtgcagcacg gtcagcacgg acaccaccct gtaggtgctg
     1081 ttgtactgct cctctctggg cttggtcttg gcgttgtgca cctccacgcc gtccacgtac
     1141 cagttgaact tcacctctgg gtcctcgtgg gacacggcca ccaccacgca ggtcacctcg
     1201 ggggtcctgc tgatcatcag ggtgtccttg ggcttggggg ggaacaggaa cacggaaggc
     1261 cctcccagca gttctggagc tgggcagggg gggcaggtgt gggtcttgtc gccgcccggc
     1321 gcgccagaac atggagaggc gggtgtgcct ggagagcttg tcactgtctc ttccacggca
     1381 ggcacttcgc cagagtgctt tgtgccgctc tctttgtgca cgcactcgat atcgctccaa
     1441 ggggtgcaat cgcccacttt gaccatgcct ctgggacagc ctgttctaca cttccggcac
     1501 atctcggggc tgtcctcttc tctgaaggtg ccttcctcgc actggcacac ggtatttctg
     1561 gtggtggtgc aagggctcag ttccacctcg ccgctatcgc atctggtgca tctcaggcag
     1621 aacagcagat cgttccagtg ggtgctgtag tcctggccgt atttgcagct gatgcagtct
     1681 ctgccgtcct cagagatgtg gtgtccagga ggacacaggc cctcagaagg gctagatctc
     1741 ttctgctgtg gggcagctct ctgctgagga gccagatcct gctgtgtaat ggcggccgca
     1801 gcgaacacca ggcagaagat gtagctcagg gcaatgatgg tcttcatggt ggcgctagca
     1861 ggcgcgtttt cgatagaacc gaggtgcagt tggacctggg agtggacacc tgtggagaga
     1921 aaggcaaagt ggatgttatt ctatagtgtc acctaaatcg tatgtgtatg atacataagg
     1981 ttatgtatta attgtagccg cgttctaacg aagccaaggg ggtgggccta tagactctat
     2041 aggcggtact tacgtcactc ttggcacggg gaatccgcgt tccaatgcac cgttcccggc
     2101 cgcggaggct ggatcggtcc cggtgtcttc tatggaggtc aaaacagcgt ggatggcgtc
     2161 tccaggcgat ctgacggttc actaaacgag ctctgcttat atagacctcc caccgtacac
     2221 gcctaccgcc catttgcgtc aatggggcgg agttgttacg acattttgga aagtcccgtt
     2281 gattttggtg ccaaaacaaa ctcccattga cgtcaatggg gtggagactt ggaaatcccc
     2341 gtgagtcaaa ccgctatcca cgcccattga tgtactgcca aaaccgcatc accatggtaa
     2401 tagcgatgac taatacgtag atgtactgcc aagtaggaaa gtcccataag gtcatgtact
     2461 gggcataatg ccaggcgggc catttaccgt cattgacgtc aatagggggc gtacttggca
     2521 tatgatacac ttgatgtact gccaagtggg cagtttaccg taaatactcc acccattgac
     2581 gtcaatggaa agtccctatt ggcgttacta tgggaacata cgtcattatt gacgtcaatg
     2641 ggcgggggtc gttgggcggt cagccaggcg ggccatttac cgtaagttat gtaacgcgga
     2701 actccatata tgggctatga actaatgacc ccgtaattga ttactattaa taactagtca
     2761 ataatcaatg ccaacatggc ggtcatattg gacatgagcc aatataaatg tacatattat
     2821 gatatagata caacgtatgc aatggccaat agccaatatt gatttatgct atataaccaa
     2881 tgaataatat ggctaatggc caatattgag tcaacgccac tgtgcatgaa gctcgtaatt
     2941 gttatccgct cacaattaag ggcgacacaa aatttattct aaatgataat aaatactgat
     3001 aacatcttat agtttgtatt atattttgta ttatcgttga catgtataat tttgatatca
     3061 aaaactgatt ttccctttat tattttcgag atttattttc ttaattctct ttaacaaact
     3121 agaaatattg tatatacaaa aaatcataaa taatagatga atagtttaat tataggtgtt
     3181 catcaatcga aaaagcaacg tatcttattt aaagtgcgtt gcttttttct catttataag
     3241 gttaaataat tctcatatat caagcaaagt gacaggcgcc cttaaatatt ctgacaaatg
     3301 ctctttccct aaactccccc cataaaaaaa cccgccgaag cgggttttta cgttatttgc
     3361 ggattaacga ttactcgtta tcagaaccgc ccagggggcc cgagcttaag actggccgtc
     3421 gttttacaac acagaaagag tttgtagaaa cgcaaaaagg ccatccgtca ggggccttct
     3481 gcttagtttg atgcctggca gttccctact ctcgccttcc gcttcctcgc tcactgactc
     3541 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     3601 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     3661 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     3721 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     3781 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     3841 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     3901 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     3961 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     4021 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     4081 tgtaggcggt gctacagagt tcttgaagtg gtgggctaac tacggctaca ctagaagaac
     4141 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     4201 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     4261 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     4321 tcagtggaac gacgcgcgcg taactcacgt taagggattt tggtcatgag caaggggtgt
     4381 tatgagccat attcaggtat aaatgggctc gcgataatgt tcagaattgg ttaattggtt
     4441 gtaacactga cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     4501 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagaatat gagtattcaa
     4561 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
     4621 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
     4681 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
     4741 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
     4801 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
     4861 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
     4921 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     4981 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     5041 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcgatg
     5101 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     5161 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     5221 gctggctggt ttattgctga taaatccgga gccggtgagc gtggttctcg cggtatcatc
     5281 gcagcgctgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     5341 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     5401 cattggtaat gaccatgatt acgccaagct ctagctagag gtcgacggta tacagacatg
     5461 ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt
     5521 atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa
     5581 gttggggtgg gcgaagaact ccagcatgag atccccgcgc tggaggatca tccagccggc
     5641 gtcccggaaa acgattccga agcccaacct ttcatagaag gcggcggtgg aatcgaaatc
     5701 tcgtagcacg tgtcaggcac cgggcttgcg ggtcatgcac caggtgcgcg gtccttcggg
     5761 cacctcgacg tcggcggtga cggtgaagcc gagccgctcg tagaagggga ggttgcgggg
     5821 cgcggatgtc tccaggaagg cgggcacccc ggcgcgctcg gccgcctcca ctccggggag
     5881 cacgacggcg ctgcccagac ccttgccctg gtggtcgggc gagacgccga cggtggccag
     5941 gaaccacgcg ggctccttgg gccggtgcgg cgccaggagg ccttccatct gttgctgcgc
     6001 ggccagccgg gaaccgctca actcggccat gcgcgggccg atctcggcga acaccgcccc
     6061 cgcttcgacg ctctccggcg tggtccagac cgccaccgcg gcgccgtcgt ccgcgaccca
     6121 caccttgccg atgtcgagcc cgacgcgcgt gaggaagagt tcttgcagct cggtgacccg
     6181 ctcgatgtgg cggtccggat cgacggtgtg gcgcgtggcg gggtagtcgg cgaacgcggc
     6241 ggcgagggtg cgtacggccc tggggacgtc gtcgcgggtg gcgaggcgca ccgtgggctt
     6301 gtactcggtc atggtttagt tcctcacctt gtcgtattat actatgccga tatactatgc
     6361 cgatgattaa ttgtcaacac gtgctgatca gatccgaaaa tggatataca agctcccggg
     6421 agctttttgc aaaagcctag gcctccaaaa aagcctcctc actacttctg gaatagctca
     6481 gaggcagagg cggcctcggc ctctgcataa ataaaaaaaa ttagtcagcc atggggcgga
     6541 gaatgggcgg aactgggcgg agttaggggc g
//
