LOCUS       sequence-368523-        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-CD70-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6571
                     /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(1930..1949)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        complement(1962..1980)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1963..1980)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2190..2214)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        complement(2211..2414)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2240..2260)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2415..2794
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     protein_bind    2961..2977
                     /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(3433..3450)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3433..3449)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3548..3565)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3719..4307)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3799..3818)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4393..4412)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(4418..4437)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        4479..4570
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4571..4639,4640..5431)
                     /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(4789..4808)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     polyA_signal    5484..5605
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5495..5514)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5549..5568
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(5735..6334)
                     /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(5818..5838)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     6315..6334
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(6353..6400)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(join(6448..6571,1..206))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(6462..6571,1..26)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(6516..6535)
                     /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 gcgcgccagg tcggacccac tgcacgccga agaatgtctc
     1381 gtcggtgttt ctgctaggca gcagggtgcc tgtcagattg gtacacaggg tatcgcctct
     1441 ggccagtggt gtcagtctct ggctggcaat tgtacagccc tggtggaagc tcagccgcag
     1501 cagagagata gatctgctgg cagggctaca gattcccacg gccagtgttg taggatggtg
     1561 tctgctggct gtggtgctgc tacagatggc cagggtcact tggatgtgca ccatgtagat
     1621 gccgtcgcgg tggattctca gctggccctt atccagctca ggtccgtgca gaaagcttct
     1681 gcccagagca ggtccacctt gccaatacag tctggggtcc tgctgaggtc ctgtgtggtt
     1741 cagctgcagt tcggcaacat cccatccaag agattccagg ggcagctgtt gctgagcctg
     1801 agcaaatctt tgggcggccg cagcgaacac caggcagaag atgtagctca gggcaatgat
     1861 ggtcttcatg gtggcgctag caggcgcgtt ttcgatagaa ccgaggtgca gttggacctg
     1921 ggagtggaca cctgtggaga gaaaggcaaa gtggatgtta ttctatagtg tcacctaaat
     1981 cgtatgtgta tgatacataa ggttatgtat taattgtagc cgcgttctaa cgaagccaag
     2041 ggggtgggcc tatagactct ataggcggta cttacgtcac tcttggcacg gggaatccgc
     2101 gttccaatgc accgttcccg gccgcggagg ctggatcggt cccggtgtct tctatggagg
     2161 tcaaaacagc gtggatggcg tctccaggcg atctgacggt tcactaaacg agctctgctt
     2221 atatagacct cccaccgtac acgcctaccg cccatttgcg tcaatggggc ggagttgtta
     2281 cgacattttg gaaagtcccg ttgattttgg tgccaaaaca aactcccatt gacgtcaatg
     2341 gggtggagac ttggaaatcc ccgtgagtca aaccgctatc cacgcccatt gatgtactgc
     2401 caaaaccgca tcaccatggt aatagcgatg actaatacgt agatgtactg ccaagtagga
     2461 aagtcccata aggtcatgta ctgggcataa tgccaggcgg gccatttacc gtcattgacg
     2521 tcaatagggg gcgtacttgg catatgatac acttgatgta ctgccaagtg ggcagtttac
     2581 cgtaaatact ccacccattg acgtcaatgg aaagtcccta ttggcgttac tatgggaaca
     2641 tacgtcatta ttgacgtcaa tgggcggggg tcgttgggcg gtcagccagg cgggccattt
     2701 accgtaagtt atgtaacgcg gaactccata tatgggctat gaactaatga ccccgtaatt
     2761 gattactatt aataactagt caataatcaa tgccaacatg gcggtcatat tggacatgag
     2821 ccaatataaa tgtacatatt atgatataga tacaacgtat gcaatggcca atagccaata
     2881 ttgatttatg ctatataacc aatgaataat atggctaatg gccaatattg agtcaacgcc
     2941 actgtgcatg aagctcgtaa ttgttatccg ctcacaatta agggcgacac aaaatttatt
     3001 ctaaatgata ataaatactg ataacatctt atagtttgta ttatattttg tattatcgtt
     3061 gacatgtata attttgatat caaaaactga ttttcccttt attattttcg agatttattt
     3121 tcttaattct ctttaacaaa ctagaaatat tgtatataca aaaaatcata aataatagat
     3181 gaatagttta attataggtg ttcatcaatc gaaaaagcaa cgtatcttat ttaaagtgcg
     3241 ttgctttttt ctcatttata aggttaaata attctcatat atcaagcaaa gtgacaggcg
     3301 cccttaaata ttctgacaaa tgctctttcc ctaaactccc cccataaaaa aacccgccga
     3361 agcgggtttt tacgttattt gcggattaac gattactcgt tatcagaacc gcccaggggg
     3421 cccgagctta agactggccg tcgttttaca acacagaaag agtttgtaga aacgcaaaaa
     3481 ggccatccgt caggggcctt ctgcttagtt tgatgcctgg cagttcccta ctctcgcctt
     3541 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     3601 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     3661 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     3721 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     3781 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     3841 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     3901 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     3961 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     4021 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     4081 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtgggcta
     4141 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     4201 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     4261 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     4321 tcttttctac ggggtctgac gctcagtgga acgacgcgcg cgtaactcac gttaagggat
     4381 tttggtcatg agcaaggggt gttatgagcc atattcaggt ataaatgggc tcgcgataat
     4441 gttcagaatt ggttaattgg ttgtaacact gacccctatt tgtttatttt tctaaataca
     4501 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     4561 aaggaagaat atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     4621 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     4681 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
     4741 ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc
     4801 ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
     4861 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
     4921 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     4981 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     5041 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     5101 caccacgatg cctgtagcga tggcaacaac gttgcgcaaa ctattaactg gcgaactact
     5161 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
     5221 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatccg gagccggtga
     5281 gcgtggttct cgcggtatca tcgcagcgct ggggccagat ggtaagccct cccgtatcgt
     5341 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     5401 gataggtgcc tcactgatta agcattggta atgaccatga ttacgccaag ctctagctag
     5461 aggtcgacgg tatacagaca tgataagata cattgatgag tttggacaaa ccacaactag
     5521 aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac
     5581 cattataagc tgcaataaac aagttggggt gggcgaagaa ctccagcatg agatccccgc
     5641 gctggaggat catccagccg gcgtcccgga aaacgattcc gaagcccaac ctttcataga
     5701 aggcggcggt ggaatcgaaa tctcgtagca cgtgtcaggc accgggcttg cgggtcatgc
     5761 accaggtgcg cggtccttcg ggcacctcga cgtcggcggt gacggtgaag ccgagccgct
     5821 cgtagaaggg gaggttgcgg ggcgcggatg tctccaggaa ggcgggcacc ccggcgcgct
     5881 cggccgcctc cactccgggg agcacgacgg cgctgcccag acccttgccc tggtggtcgg
     5941 gcgagacgcc gacggtggcc aggaaccacg cgggctcctt gggccggtgc ggcgccagga
     6001 ggccttccat ctgttgctgc gcggccagcc gggaaccgct caactcggcc atgcgcgggc
     6061 cgatctcggc gaacaccgcc cccgcttcga cgctctccgg cgtggtccag accgccaccg
     6121 cggcgccgtc gtccgcgacc cacaccttgc cgatgtcgag cccgacgcgc gtgaggaaga
     6181 gttcttgcag ctcggtgacc cgctcgatgt ggcggtccgg atcgacggtg tggcgcgtgg
     6241 cggggtagtc ggcgaacgcg gcggcgaggg tgcgtacggc cctggggacg tcgtcgcggg
     6301 tggcgaggcg caccgtgggc ttgtactcgg tcatggttta gttcctcacc ttgtcgtatt
     6361 atactatgcc gatatactat gccgatgatt aattgtcaac acgtgctgat cagatccgaa
     6421 aatggatata caagctcccg ggagcttttt gcaaaagcct aggcctccaa aaaagcctcc
     6481 tcactacttc tggaatagct cagaggcaga ggcggcctcg gcctctgcat aaataaaaaa
     6541 aattagtcag ccatggggcg gagaatgggc g
//
