LOCUS       sequence-368743-        6625 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 6625)
  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 6625)
  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-CD40-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6625
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     194..214
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    332..443
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     432..449
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(589..606)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(607..651)
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     primer_bind     complement(1987..2006)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        complement(2019..2037)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2020..2037)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2247..2271)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        complement(2268..2471)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2297..2317)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2472..2851
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     protein_bind    3018..3034
                     /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(3490..3507)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3490..3506)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3605..3622)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3776..4364)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3856..3875)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4450..4469)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(4475..4494)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        4536..4627
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4628..4696,4697..5488)
                     /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(4846..4865)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     polyA_signal    5541..5662
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5552..5571)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5606..5625
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(5792..6391)
                     /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(5875..5895)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     6372..6391
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(6410..6457)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(join(6505..6625,1..209))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(6519..6625,1..29)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(6573..6592)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
ORIGIN
        1 gcggaactgg gcggagttag gggcgggatg ggcggagtta ggggcgggac tatggttgct
       61 gactaattga gatgcatgct ttgcatactt ctgcctgctg gggagcctgg ggactttcca
      121 cacctggttg ctgactaatt gagatgcatg ctttgcatac ttctgcctgc tggggagcct
      181 ggggactttc cacaccctaa ctgacacaca ttccacagaa ttaattcgcg ttaaattttt
      241 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
      301 aagaatagac cgagataggg ttgagtgttg tgatgcaatt tcctcatttt attaggaaag
      361 gacagtggga gtggcacctt ccagggtcaa ggaaggcacg ggggaggggc aaacaacaga
      421 tggctggcaa ctagaaggca cagtcgaggc tgattttctc tatctattta ttttactact
      481 caagactatc ttgcaaagta gaagaaccct gtctgcagta tgacgaccgt gtatacccga
      541 ttggcttacc gggtaacgcg ttgtgccagt gactacaggc cggccttagt ggtgatggtg
      601 gtggtgttca tgccactcaa tcttctgagc ctcgaaaata tcattaagac cgctgccgcc
      661 ggggctcagg ctcagggact tctgggtgta gtggttgtgc agggcctcgt gcatcacgct
      721 gcagctgaac acgttgccct gctgccacct ggacttgtcc acggtcagct tgctgtacag
      781 gaagaagctg ccgtcgctgt ccagcactgg gggggtggtc ttgtagttgt tctcgggctg
      841 gccgttgctc tcccactcca cggcgatatc gctggggtag aagcccttca ccagacaggt
      901 cagggacacc tggttcttgg tcatctcctc ccggctgggg ggcagggtgt acacctgggg
      961 ctcccgtggc tggcccttgg ccttgctgat tgtcttttcg attggggcag ccagggcctt
     1021 gttggagact ttgcacttgt attctttgcc gttcagccag tcctggtgca gcacggtcag
     1081 cacggacacc accctgtagg tgctgttgta ctgctcctct ctgggcttgg tcttggcgtt
     1141 gtgcacctcc acgccgtcca cgtaccagtt gaacttcacc tctgggtcct cgtgggacac
     1201 ggccaccacc acgcaggtca cctcgggggt cctgctgatc atcagggtgt ccttgggctt
     1261 gggggggaac aggaacacgg aaggccctcc cagcagttct ggagctgggc agggggggca
     1321 ggtgtgggtc ttgtcgccgc ccggcgcgcc tctaagccgg tcctgagggc cgcacacaac
     1381 atctgtcttg ttggtgccgg cctgctgcac aaccagatcc tttgtctcgc agctggtcca
     1441 agggtgacac ttctcgaagg cgctggacac attgctgaag aagcccacag gacaaggctc
     1501 gcagatggta tcggacacgc ctgtggcaat ctgcttcact ccgaagccag ggctacagct
     1561 tctgtgcagc acacagcttt cacaggcctc gctggtacag tgccatcctt cctcgcatgt
     1621 acagatggtg tctgtctcgc tggtgccctt ctgctgcact ctcagtccca gattggggtc
     1681 gcagtacttg tgctggtggc agtgtgtctc tctattccag gtgtccagga actcgctctc
     1741 gccgcaaggc agacactctg tctcggtgaa ctcggtgcaa tcggacacca gtttctgtcc
     1801 aggctggcac aggctgcagc actggctgtt gatcaggtac tgcttctctc tacaggctgt
     1861 aggaggttcg gcggccgcag cgaacaccag gcagaagatg tagctcaggg caatgatggt
     1921 cttcatggtg gcgctagcag gcgcgttttc gatagaaccg aggtgcagtt ggacctggga
     1981 gtggacacct gtggagagaa aggcaaagtg gatgttattc tatagtgtca cctaaatcgt
     2041 atgtgtatga tacataaggt tatgtattaa ttgtagccgc gttctaacga agccaagggg
     2101 gtgggcctat agactctata ggcggtactt acgtcactct tggcacgggg aatccgcgtt
     2161 ccaatgcacc gttcccggcc gcggaggctg gatcggtccc ggtgtcttct atggaggtca
     2221 aaacagcgtg gatggcgtct ccaggcgatc tgacggttca ctaaacgagc tctgcttata
     2281 tagacctccc accgtacacg cctaccgccc atttgcgtca atggggcgga gttgttacga
     2341 cattttggaa agtcccgttg attttggtgc caaaacaaac tcccattgac gtcaatgggg
     2401 tggagacttg gaaatccccg tgagtcaaac cgctatccac gcccattgat gtactgccaa
     2461 aaccgcatca ccatggtaat agcgatgact aatacgtaga tgtactgcca agtaggaaag
     2521 tcccataagg tcatgtactg ggcataatgc caggcgggcc atttaccgtc attgacgtca
     2581 atagggggcg tacttggcat atgatacact tgatgtactg ccaagtgggc agtttaccgt
     2641 aaatactcca cccattgacg tcaatggaaa gtccctattg gcgttactat gggaacatac
     2701 gtcattattg acgtcaatgg gcgggggtcg ttgggcggtc agccaggcgg gccatttacc
     2761 gtaagttatg taacgcggaa ctccatatat gggctatgaa ctaatgaccc cgtaattgat
     2821 tactattaat aactagtcaa taatcaatgc caacatggcg gtcatattgg acatgagcca
     2881 atataaatgt acatattatg atatagatac aacgtatgca atggccaata gccaatattg
     2941 atttatgcta tataaccaat gaataatatg gctaatggcc aatattgagt caacgccact
     3001 gtgcatgaag ctcgtaattg ttatccgctc acaattaagg gcgacacaaa atttattcta
     3061 aatgataata aatactgata acatcttata gtttgtatta tattttgtat tatcgttgac
     3121 atgtataatt ttgatatcaa aaactgattt tccctttatt attttcgaga tttattttct
     3181 taattctctt taacaaacta gaaatattgt atatacaaaa aatcataaat aatagatgaa
     3241 tagtttaatt ataggtgttc atcaatcgaa aaagcaacgt atcttattta aagtgcgttg
     3301 cttttttctc atttataagg ttaaataatt ctcatatatc aagcaaagtg acaggcgccc
     3361 ttaaatattc tgacaaatgc tctttcccta aactcccccc ataaaaaaac ccgccgaagc
     3421 gggtttttac gttatttgcg gattaacgat tactcgttat cagaaccgcc cagggggccc
     3481 gagcttaaga ctggccgtcg ttttacaaca cagaaagagt ttgtagaaac gcaaaaaggc
     3541 catccgtcag gggccttctg cttagtttga tgcctggcag ttccctactc tcgccttccg
     3601 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     3661 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     3721 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     3781 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     3841 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     3901 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     3961 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     4021 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     4081 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     4141 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tgggctaact
     4201 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     4261 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     4321 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     4381 tttctacggg gtctgacgct cagtggaacg acgcgcgcgt aactcacgtt aagggatttt
     4441 ggtcatgagc aaggggtgtt atgagccata ttcaggtata aatgggctcg cgataatgtt
     4501 cagaattggt taattggttg taacactgac ccctatttgt ttatttttct aaatacattc
     4561 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     4621 gaagaatatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg
     4681 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt
     4741 gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt
     4801 tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt
     4861 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa
     4921 tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag
     4981 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac
     5041 aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac
     5101 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
     5161 cacgatgcct gtagcgatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac
     5221 tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact
     5281 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatccggag ccggtgagcg
     5341 tggttctcgc ggtatcatcg cagcgctggg gccagatggt aagccctccc gtatcgtagt
     5401 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
     5461 aggtgcctca ctgattaagc attggtaatg accatgatta cgccaagctc tagctagagg
     5521 tcgacggtat acagacatga taagatacat tgatgagttt ggacaaacca caactagaat
     5581 gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat
     5641 tataagctgc aataaacaag ttggggtggg cgaagaactc cagcatgaga tccccgcgct
     5701 ggaggatcat ccagccggcg tcccggaaaa cgattccgaa gcccaacctt tcatagaagg
     5761 cggcggtgga atcgaaatct cgtagcacgt gtcaggcacc gggcttgcgg gtcatgcacc
     5821 aggtgcgcgg tccttcgggc acctcgacgt cggcggtgac ggtgaagccg agccgctcgt
     5881 agaaggggag gttgcggggc gcggatgtct ccaggaaggc gggcaccccg gcgcgctcgg
     5941 ccgcctccac tccggggagc acgacggcgc tgcccagacc cttgccctgg tggtcgggcg
     6001 agacgccgac ggtggccagg aaccacgcgg gctccttggg ccggtgcggc gccaggaggc
     6061 cttccatctg ttgctgcgcg gccagccggg aaccgctcaa ctcggccatg cgcgggccga
     6121 tctcggcgaa caccgccccc gcttcgacgc tctccggcgt ggtccagacc gccaccgcgg
     6181 cgccgtcgtc cgcgacccac accttgccga tgtcgagccc gacgcgcgtg aggaagagtt
     6241 cttgcagctc ggtgacccgc tcgatgtggc ggtccggatc gacggtgtgg cgcgtggcgg
     6301 ggtagtcggc gaacgcggcg gcgagggtgc gtacggccct ggggacgtcg tcgcgggtgg
     6361 cgaggcgcac cgtgggcttg tactcggtca tggtttagtt cctcaccttg tcgtattata
     6421 ctatgccgat atactatgcc gatgattaat tgtcaacacg tgctgatcag atccgaaaat
     6481 ggatatacaa gctcccggga gctttttgca aaagcctagg cctccaaaaa agcctcctca
     6541 ctacttctgg aatagctcag aggcagaggc ggcctcggcc tctgcataaa taaaaaaaat
     6601 tagtcagcca tggggcggag aatgg
//
