LOCUS       sequence-368525-        6574 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 6574)
  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 6574)
  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-TNFRSF10D-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6574
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     191..211
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    329..440
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     429..446
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(586..603)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(604..648)
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     primer_bind     complement(1933..1952)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        complement(1965..1983)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1966..1983)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2193..2217)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        complement(2214..2417)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2243..2263)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2418..2797
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     protein_bind    2964..2980
                     /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(3436..3453)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3436..3452)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3551..3568)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3722..4310)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3802..3821)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4396..4415)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(4421..4440)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        4482..4573
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4574..4642,4643..5434)
                     /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(4792..4811)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     polyA_signal    5487..5608
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5498..5517)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5552..5571
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(5738..6337)
                     /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(5821..5841)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     6318..6337
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(6356..6403)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(join(6451..6574,1..206))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(6465..6574,1..26)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(6519..6538)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
ORIGIN
        1 gaactgggcg gagttagggg cgggatgggc ggagttaggg gcgggactat ggttgctgac
       61 taattgagat gcatgctttg catacttctg cctgctgggg agcctgggga ctttccacac
      121 ctggttgctg actaattgag atgcatgctt tgcatacttc tgcctgctgg ggagcctggg
      181 gactttccac accctaactg acacacattc cacagaatta attcgcgtta aatttttgtt
      241 aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag
      301 aatagaccga gatagggttg agtgttgtga tgcaatttcc tcattttatt aggaaaggac
      361 agtgggagtg gcaccttcca gggtcaagga aggcacgggg gaggggcaaa caacagatgg
      421 ctggcaacta gaaggcacag tcgaggctga ttttctctat ctatttattt tactactcaa
      481 gactatcttg caaagtagaa gaaccctgtc tgcagtatga cgaccgtgta tacccgattg
      541 gcttaccggg taacgcgttg tgccagtgac tacaggccgg ccttagtggt gatggtggtg
      601 gtgttcatgc cactcaatct tctgagcctc gaaaatatca ttaagaccgc tgccgccggg
      661 gctcaggctc agggacttct gggtgtagtg gttgtgcagg gcctcgtgca tcacgctgca
      721 gctgaacacg ttgccctgct gccacctgga cttgtccacg gtcagcttgc tgtacaggaa
      781 gaagctgccg tcgctgtcca gcactggggg ggtggtcttg tagttgttct cgggctggcc
      841 gttgctctcc cactccacgg cgatatcgct ggggtagaag cccttcacca gacaggtcag
      901 ggacacctgg ttcttggtca tctcctcccg gctggggggc agggtgtaca cctggggctc
      961 ccgtggctgg cccttggcct tgctgattgt cttttcgatt ggggcagcca gggccttgtt
     1021 ggagactttg cacttgtatt ctttgccgtt cagccagtcc tggtgcagca cggtcagcac
     1081 ggacaccacc ctgtaggtgc tgttgtactg ctcctctctg ggcttggtct tggcgttgtg
     1141 cacctccacg ccgtccacgt accagttgaa cttcacctct gggtcctcgt gggacacggc
     1201 caccaccacg caggtcacct cgggggtcct gctgatcatc agggtgtcct tgggcttggg
     1261 ggggaacagg aacacggaag gccctcccag cagttctgga gctgggcagg gggggcaggt
     1321 gtgggtcttg tcgccgcccg gcgcgccatg gtaaggagag gccagcattc ccagaatggt
     1381 ggtcacggtt tcctcggcag ctggtgtttt gcctgtgctg ctggcggcgc tctcattctt
     1441 gcacttgatg tcgctccgag gggtgcaatt ggacaccttg accatgcctc tgggacagcc
     1501 tgttctgcag gttctgcaca tctcggggct gttcttgtcc tggaagctgc ccttctcgca
     1561 ctgacacacg gtatctctgg tggtggtgca gctgctcttg ttggtctggc cgctcttgca
     1621 cactgtacac agcaggcagc taggcaggtt gttgctggcg attgtgtagt ccacgccttc
     1681 ggtgcaaggg ttgcaagcgc ctgtgtactc agatctgtgg ctgccggctg ggcactcttc
     1741 ctctttcagg cttcttctct gctgctgggg agcaactgtc tgctggggca cttcatcctg
     1801 tctaggaatt gttgcggcgg ccgcagcgaa caccaggcag aagatgtagc tcagggcaat
     1861 gatggtcttc atggtggcgc tagcaggcgc gttttcgata gaaccgaggt gcagttggac
     1921 ctgggagtgg acacctgtgg agagaaaggc aaagtggatg ttattctata gtgtcaccta
     1981 aatcgtatgt gtatgataca taaggttatg tattaattgt agccgcgttc taacgaagcc
     2041 aagggggtgg gcctatagac tctataggcg gtacttacgt cactcttggc acggggaatc
     2101 cgcgttccaa tgcaccgttc ccggccgcgg aggctggatc ggtcccggtg tcttctatgg
     2161 aggtcaaaac agcgtggatg gcgtctccag gcgatctgac ggttcactaa acgagctctg
     2221 cttatataga cctcccaccg tacacgccta ccgcccattt gcgtcaatgg ggcggagttg
     2281 ttacgacatt ttggaaagtc ccgttgattt tggtgccaaa acaaactccc attgacgtca
     2341 atggggtgga gacttggaaa tccccgtgag tcaaaccgct atccacgccc attgatgtac
     2401 tgccaaaacc gcatcaccat ggtaatagcg atgactaata cgtagatgta ctgccaagta
     2461 ggaaagtccc ataaggtcat gtactgggca taatgccagg cgggccattt accgtcattg
     2521 acgtcaatag ggggcgtact tggcatatga tacacttgat gtactgccaa gtgggcagtt
     2581 taccgtaaat actccaccca ttgacgtcaa tggaaagtcc ctattggcgt tactatggga
     2641 acatacgtca ttattgacgt caatgggcgg gggtcgttgg gcggtcagcc aggcgggcca
     2701 tttaccgtaa gttatgtaac gcggaactcc atatatgggc tatgaactaa tgaccccgta
     2761 attgattact attaataact agtcaataat caatgccaac atggcggtca tattggacat
     2821 gagccaatat aaatgtacat attatgatat agatacaacg tatgcaatgg ccaatagcca
     2881 atattgattt atgctatata accaatgaat aatatggcta atggccaata ttgagtcaac
     2941 gccactgtgc atgaagctcg taattgttat ccgctcacaa ttaagggcga cacaaaattt
     3001 attctaaatg ataataaata ctgataacat cttatagttt gtattatatt ttgtattatc
     3061 gttgacatgt ataattttga tatcaaaaac tgattttccc tttattattt tcgagattta
     3121 ttttcttaat tctctttaac aaactagaaa tattgtatat acaaaaaatc ataaataata
     3181 gatgaatagt ttaattatag gtgttcatca atcgaaaaag caacgtatct tatttaaagt
     3241 gcgttgcttt tttctcattt ataaggttaa ataattctca tatatcaagc aaagtgacag
     3301 gcgcccttaa atattctgac aaatgctctt tccctaaact ccccccataa aaaaacccgc
     3361 cgaagcgggt ttttacgtta tttgcggatt aacgattact cgttatcaga accgcccagg
     3421 gggcccgagc ttaagactgg ccgtcgtttt acaacacaga aagagtttgt agaaacgcaa
     3481 aaaggccatc cgtcaggggc cttctgctta gtttgatgcc tggcagttcc ctactctcgc
     3541 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     3601 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     3661 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     3721 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     3781 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     3841 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     3901 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     3961 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     4021 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     4081 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggg
     4141 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     4201 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     4261 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     4321 tgatcttttc tacggggtct gacgctcagt ggaacgacgc gcgcgtaact cacgttaagg
     4381 gattttggtc atgagcaagg ggtgttatga gccatattca ggtataaatg ggctcgcgat
     4441 aatgttcaga attggttaat tggttgtaac actgacccct atttgtttat ttttctaaat
     4501 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     4561 aaaaaggaag aatatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     4621 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     4681 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     4741 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
     4801 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     4861 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     4921 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     4981 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     5041 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     5101 tgacaccacg atgcctgtag cgatggcaac aacgttgcgc aaactattaa ctggcgaact
     5161 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     5221 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ccggagccgg
     5281 tgagcgtggt tctcgcggta tcatcgcagc gctggggcca gatggtaagc cctcccgtat
     5341 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     5401 tgagataggt gcctcactga ttaagcattg gtaatgacca tgattacgcc aagctctagc
     5461 tagaggtcga cggtatacag acatgataag atacattgat gagtttggac aaaccacaac
     5521 tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt
     5581 aaccattata agctgcaata aacaagttgg ggtgggcgaa gaactccagc atgagatccc
     5641 cgcgctggag gatcatccag ccggcgtccc ggaaaacgat tccgaagccc aacctttcat
     5701 agaaggcggc ggtggaatcg aaatctcgta gcacgtgtca ggcaccgggc ttgcgggtca
     5761 tgcaccaggt gcgcggtcct tcgggcacct cgacgtcggc ggtgacggtg aagccgagcc
     5821 gctcgtagaa ggggaggttg cggggcgcgg atgtctccag gaaggcgggc accccggcgc
     5881 gctcggccgc ctccactccg gggagcacga cggcgctgcc cagacccttg ccctggtggt
     5941 cgggcgagac gccgacggtg gccaggaacc acgcgggctc cttgggccgg tgcggcgcca
     6001 ggaggccttc catctgttgc tgcgcggcca gccgggaacc gctcaactcg gccatgcgcg
     6061 ggccgatctc ggcgaacacc gcccccgctt cgacgctctc cggcgtggtc cagaccgcca
     6121 ccgcggcgcc gtcgtccgcg acccacacct tgccgatgtc gagcccgacg cgcgtgagga
     6181 agagttcttg cagctcggtg acccgctcga tgtggcggtc cggatcgacg gtgtggcgcg
     6241 tggcggggta gtcggcgaac gcggcggcga gggtgcgtac ggccctgggg acgtcgtcgc
     6301 gggtggcgag gcgcaccgtg ggcttgtact cggtcatggt ttagttcctc accttgtcgt
     6361 attatactat gccgatatac tatgccgatg attaattgtc aacacgtgct gatcagatcc
     6421 gaaaatggat atacaagctc ccgggagctt tttgcaaaag cctaggcctc caaaaaagcc
     6481 tcctcactac ttctggaata gctcagaggc agaggcggcc tcggcctctg cataaataaa
     6541 aaaaattagt cagccatggg gcggagaatg ggcg
//
