LOCUS       sequence-368824-        6466 bp    DNA     circular SYN 14-APR-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 6466)
  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 6466)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 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-ICOS-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6466
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     238..257
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(276..323)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(371..700)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      385..520
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(439..458)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     685..705
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    823..934
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     923..940
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(1080..1097)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(1098..1142)
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     primer_bind     complement(2319..2338)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        complement(2351..2369)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2352..2369)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2579..2603)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        complement(2600..2803)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2629..2649)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2804..3183
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     protein_bind    3350..3366
                     /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(3822..3839)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3822..3838)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3937..3954)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4108..4696)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4188..4207)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4782..4801)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(4807..4826)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        4868..4959
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4960..5028,5029..5820)
                     /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(5178..5197)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     polyA_signal    5873..5994
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5884..5903)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5938..5957
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(join(6124..6466,1..257))
                     /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(6207..6227)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
ORIGIN
        1 gcccccgctt cgacgctctc cggcgtggtc cagaccgcca ccgcggcgcc gtcgtccgcg
       61 acccacacct tgccgatgtc gagcccgacg cgcgtgagga agagttcttg cagctcggtg
      121 acccgctcga tgtggcggtc cggatcgacg gtgtggcgcg tggcggggta gtcggcgaac
      181 gcggcggcga gggtgcgtac ggccctgggg acgtcgtcgc gggtggcgag gcgcaccgtg
      241 ggcttgtact cggtcatggt ttagttcctc accttgtcgt attatactat gccgatatac
      301 tatgccgatg attaattgtc aacacgtgct gatcagatcc gaaaatggat atacaagctc
      361 ccgggagctt tttgcaaaag cctaggcctc caaaaaagcc tcctcactac ttctggaata
      421 gctcagaggc agaggcggcc tcggcctctg cataaataaa aaaaattagt cagccatggg
      481 gcggagaatg ggcggaactg ggcggagtta ggggcgggat gggcggagtt aggggcggga
      541 ctatggttgc tgactaattg agatgcatgc tttgcatact tctgcctgct ggggagcctg
      601 gggactttcc acacctggtt gctgactaat tgagatgcat gctttgcata cttctgcctg
      661 ctggggagcc tggggacttt ccacacccta actgacacac attccacaga attaattcgc
      721 gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc
      781 ttataaatca aaagaataga ccgagatagg gttgagtgtt gtgatgcaat ttcctcattt
      841 tattaggaaa ggacagtggg agtggcacct tccagggtca aggaaggcac gggggagggg
      901 caaacaacag atggctggca actagaaggc acagtcgagg ctgattttct ctatctattt
      961 attttactac tcaagactat cttgcaaagt agaagaaccc tgtctgcagt atgacgaccg
     1021 tgtatacccg attggcttac cgggtaacgc gttgtgccag tgactacagg ccggccttag
     1081 tggtgatggt ggtggtgttc atgccactca atcttctgag cctcgaaaat atcattaaga
     1141 ccgctgccgc cggggctcag gctcagggac ttctgggtgt agtggttgtg cagggcctcg
     1201 tgcatcacgc tgcagctgaa cacgttgccc tgctgccacc tggacttgtc cacggtcagc
     1261 ttgctgtaca ggaagaagct gccgtcgctg tccagcactg ggggggtggt cttgtagttg
     1321 ttctcgggct ggccgttgct ctcccactcc acggcgatat cgctggggta gaagcccttc
     1381 accagacagg tcagggacac ctggttcttg gtcatctcct cccggctggg gggcagggtg
     1441 tacacctggg gctcccgtgg ctggcccttg gccttgctga ttgtcttttc gattggggca
     1501 gccagggcct tgttggagac tttgcacttg tattctttgc cgttcagcca gtcctggtgc
     1561 agcacggtca gcacggacac caccctgtag gtgctgttgt actgctcctc tctgggcttg
     1621 gtcttggcgt tgtgcacctc cacgccgtcc acgtaccagt tgaacttcac ctctgggtcc
     1681 tcgtgggaca cggccaccac cacgcaggtc acctcggggg tcctgctgat catcagggtg
     1741 tccttgggct tgggggggaa caggaacacg gaaggccctc ccagcagttc tggagctggg
     1801 cagggggggc aggtgtgggt cttgtcgccg cccggcgcgc ctttcagctg gcagcacagc
     1861 tggctctcgt agatgtgcag gtagccgcct gtcagggtca ctttgaatgg aggcgggtcg
     1921 aagatgctca ggttgcagaa atagtaattg gcgtggctgt ggtccaggtt gtacaggaag
     1981 aaggacacgc tgttgttgct cagctggctg tggcagaact tcaggctctt gatggacacg
     2041 gtgttgccgc tgccttttgt cttggtcagg tcgcacagaa tctggccgcc tttcagcagc
     2101 tgcatcttga actgctgcac gatatcaggg tacttgcaca ggatctgcac gccgccgttg
     2161 tggaagatga acatctcgta gttggcgctg ccattgattt cggcggccgc agcgaacacc
     2221 aggcagaaga tgtagctcag ggcaatgatg gtcttcatgg tggcgctagc aggcgcgttt
     2281 tcgatagaac cgaggtgcag ttggacctgg gagtggacac ctgtggagag aaaggcaaag
     2341 tggatgttat tctatagtgt cacctaaatc gtatgtgtat gatacataag gttatgtatt
     2401 aattgtagcc gcgttctaac gaagccaagg gggtgggcct atagactcta taggcggtac
     2461 ttacgtcact cttggcacgg ggaatccgcg ttccaatgca ccgttcccgg ccgcggaggc
     2521 tggatcggtc ccggtgtctt ctatggaggt caaaacagcg tggatggcgt ctccaggcga
     2581 tctgacggtt cactaaacga gctctgctta tatagacctc ccaccgtaca cgcctaccgc
     2641 ccatttgcgt caatggggcg gagttgttac gacattttgg aaagtcccgt tgattttggt
     2701 gccaaaacaa actcccattg acgtcaatgg ggtggagact tggaaatccc cgtgagtcaa
     2761 accgctatcc acgcccattg atgtactgcc aaaaccgcat caccatggta atagcgatga
     2821 ctaatacgta gatgtactgc caagtaggaa agtcccataa ggtcatgtac tgggcataat
     2881 gccaggcggg ccatttaccg tcattgacgt caataggggg cgtacttggc atatgataca
     2941 cttgatgtac tgccaagtgg gcagtttacc gtaaatactc cacccattga cgtcaatgga
     3001 aagtccctat tggcgttact atgggaacat acgtcattat tgacgtcaat gggcgggggt
     3061 cgttgggcgg tcagccaggc gggccattta ccgtaagtta tgtaacgcgg aactccatat
     3121 atgggctatg aactaatgac cccgtaattg attactatta ataactagtc aataatcaat
     3181 gccaacatgg cggtcatatt ggacatgagc caatataaat gtacatatta tgatatagat
     3241 acaacgtatg caatggccaa tagccaatat tgatttatgc tatataacca atgaataata
     3301 tggctaatgg ccaatattga gtcaacgcca ctgtgcatga agctcgtaat tgttatccgc
     3361 tcacaattaa gggcgacaca aaatttattc taaatgataa taaatactga taacatctta
     3421 tagtttgtat tatattttgt attatcgttg acatgtataa ttttgatatc aaaaactgat
     3481 tttcccttta ttattttcga gatttatttt cttaattctc tttaacaaac tagaaatatt
     3541 gtatatacaa aaaatcataa ataatagatg aatagtttaa ttataggtgt tcatcaatcg
     3601 aaaaagcaac gtatcttatt taaagtgcgt tgcttttttc tcatttataa ggttaaataa
     3661 ttctcatata tcaagcaaag tgacaggcgc ccttaaatat tctgacaaat gctctttccc
     3721 taaactcccc ccataaaaaa acccgccgaa gcgggttttt acgttatttg cggattaacg
     3781 attactcgtt atcagaaccg cccagggggc ccgagcttaa gactggccgt cgttttacaa
     3841 cacagaaaga gtttgtagaa acgcaaaaag gccatccgtc aggggccttc tgcttagttt
     3901 gatgcctggc agttccctac tctcgccttc cgcttcctcg ctcactgact cgctgcgctc
     3961 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     4021 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     4081 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     4141 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     4201 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     4261 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     4321 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     4381 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     4441 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     4501 tgctacagag ttcttgaagt ggtgggctaa ctacggctac actagaagaa cagtatttgg
     4561 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     4621 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     4681 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     4741 cgacgcgcgc gtaactcacg ttaagggatt ttggtcatga gcaaggggtg ttatgagcca
     4801 tattcaggta taaatgggct cgcgataatg ttcagaattg gttaattggt tgtaacactg
     4861 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     4921 ccctgataaa tgcttcaata atattgaaaa aggaagaata tgagtattca acatttccgt
     4981 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     5041 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     5101 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     5161 agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag
     5221 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     5281 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     5341 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     5401 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     5461 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcgat ggcaacaacg
     5521 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     5581 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     5641 tttattgctg ataaatccgg agccggtgag cgtggttctc gcggtatcat cgcagcgctg
     5701 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     5761 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     5821 tgaccatgat tacgccaagc tctagctaga ggtcgacggt atacagacat gataagatac
     5881 attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa
     5941 atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca agttggggtg
     6001 ggcgaagaac tccagcatga gatccccgcg ctggaggatc atccagccgg cgtcccggaa
     6061 aacgattccg aagcccaacc tttcatagaa ggcggcggtg gaatcgaaat ctcgtagcac
     6121 gtgtcaggca ccgggcttgc gggtcatgca ccaggtgcgc ggtccttcgg gcacctcgac
     6181 gtcggcggtg acggtgaagc cgagccgctc gtagaagggg aggttgcggg gcgcggatgt
     6241 ctccaggaag gcgggcaccc cggcgcgctc ggccgcctcc actccgggga gcacgacggc
     6301 gctgcccaga cccttgccct ggtggtcggg cgagacgccg acggtggcca ggaaccacgc
     6361 gggctccttg ggccggtgcg gcgccaggag gccttccatc tgttgctgcg cggccagccg
     6421 ggaaccgctc aactcggcca tgcgcgggcc gatctcggcg aacacc
//
