LOCUS       sequence-430454-        6769 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 6769)
  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 6769)
  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-KIR2DL4-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6769
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        77..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     581..605
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        815..833
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     815..832
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     846..865
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             2345..2389
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             2390..2407
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2547..2564)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2553..2664
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2782..2802)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2787..3116
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2967..3102
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3029..3048
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3164..3211
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3230..3829
                     /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(3230..3249)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3726..3746
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    3959..4080
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3996..4015)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4050..4069
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(4133..4924,4925..4993))
                     /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     4756..4775
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4994..5085)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5127..5146
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5152..5171
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      5257..5845
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5746..5765
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5999..6016
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     6114..6131
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6115..6131
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6587..6603
                     /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)."
ORIGIN
        1 ggcattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
      601 agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagacac cgggaccgat
      661 ccagcctccg cggccgggaa cggtgcattg gaacgcggat tccccgtgcc aagagtgacg
      721 taagtaccgc ctatagagtc tataggccca cccccttggc ttcgttagaa cgcggctaca
      781 attaatacat aaccttatgt atcatacaca tacgatttag gtgacactat agaataacat
      841 ccactttgcc tttctctcca caggtgtcca ctcccaggtc caactgcacc tcggttctat
      901 cgaaaacgcg cctgctagcg ccaccatgaa gaccatcatt gccctgagct acatcttctg
      961 cctggtgttc gctgcggccg cctgggctca tgttggagga caggataagc ctttctgcag
     1021 cgcctggcct tctgctgttg ttcctcaagg cggacacgtg accctgcggt gtcactacag
     1081 acggggcttc aacatcttca ccctgtacaa gaaagacggc gtgcccgtgc ctgagctgta
     1141 caacagaatc ttctggaaca gcttcctgat cagccccgtg acaccagctc acgccggcac
     1201 atacagatgc agaggctttc accctcacag ccccacagag tggtccgctc catctaaccc
     1261 tctggtcatc atggtcaccg gcctgtacga gaagcctagc ctgacagcta gacccggacc
     1321 tacagttaga gccggcgaga atgtgacact gagctgcagc agccagagca gcttcgacat
     1381 ctaccacctg agcagagagg gcgaagccca cgaacttaga ctgcctgccg tgcctagcat
     1441 caacggcacc tttcaggccg attttccact gggacctgcc acacacggcg agacttacag
     1501 atgctttggc agcttccacg gcagccctta cgagtggtct gatcctagcg atcctctgcc
     1561 tgtgtccgtg acaggcaatc ctagcagcag ctggccatct cctaccgagc ctagctttaa
     1621 gaccggaatc gcccggcatc tgcatggcgc gccgggcggc gacaagaccc acacctgccc
     1681 cccctgccca gctccagaac tgctgggagg gccttccgtg ttcctgttcc cccccaagcc
     1741 caaggacacc ctgatgatca gcaggacccc cgaggtgacc tgcgtggtgg tggccgtgtc
     1801 ccacgaggac ccagaggtga agttcaactg gtacgtggac ggcgtggagg tgcacaacgc
     1861 caagaccaag cccagagagg agcagtacaa cagcacctac agggtggtgt ccgtgctgac
     1921 cgtgctgcac caggactggc tgaacggcaa agaatacaag tgcaaagtct ccaacaaggc
     1981 cctggctgcc ccaatcgaaa agacaatcag caaggccaag ggccagccac gggagcccca
     2041 ggtgtacacc ctgcccccca gccgggagga gatgaccaag aaccaggtgt ccctgacctg
     2101 tctggtgaag ggcttctacc ccagcgatat cgccgtggag tgggagagca acggccagcc
     2161 cgagaacaac tacaagacca cccccccagt gctggacagc gacggcagct tcttcctgta
     2221 cagcaagctg accgtggaca agtccaggtg gcagcagggc aacgtgttca gctgcagcgt
     2281 gatgcacgag gccctgcaca accactacac ccagaagtcc ctgagcctga gccccggcgg
     2341 cagcggtctt aatgatattt tcgaggctca gaagattgag tggcatgaac accaccacca
     2401 tcaccactaa ggccggcctg tagtcactgg cacaacgcgt tacccggtaa gccaatcggg
     2461 tatacacggt cgtcatactg cagacagggt tcttctactt tgcaagatag tcttgagtag
     2521 taaaataaat agatagagaa aatcagcctc gactgtgcct tctagttgcc agccatctgt
     2581 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     2641 ctaataaaat gaggaaattg catcacaaca ctcaacccta tctcggtcta ttcttttgat
     2701 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     2761 tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
     2821 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
     2881 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     2941 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     3001 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct
     3061 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
     3121 tcccgggagc ttgtatatcc attttcggat ctgatcagca cgtgttgaca attaatcatc
     3181 ggcatagtat atcggcatag tataatacga caaggtgagg aactaaacca tgaccgagta
     3241 caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac gcaccctcgc
     3301 cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc gatccggacc gccacatcga
     3361 gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt
     3421 gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc
     3481 gggggcggtg ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc
     3541 cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt
     3601 cctggccacc gtcggcgtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt
     3661 gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg agacatccgc
     3721 gccccgcaac ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt
     3781 gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgac acgtgctacg
     3841 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga
     3901 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa
     3961 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     4021 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     4081 tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt cattaccaat
     4141 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     4201 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagcgctg
     4261 cgatgatacc gcgagaacca cgctcaccgg ctccggattt atcagcaata aaccagccag
     4321 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     4381 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     4441 ccatcgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     4501 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     4561 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     4621 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     4681 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     4741 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     4801 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     4861 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     4921 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     4981 gttgaatact catattcttc ctttttcaat attattgaag catttatcag ggttattgtc
     5041 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gtcagtgtta
     5101 caaccaatta accaattctg aacattatcg cgagcccatt tatacctgaa tatggctcat
     5161 aacacccctt gctcatgacc aaaatccctt aacgtgagtt acgcgcgcgt cgttccactg
     5221 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     5281 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     5341 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     5401 tgttcttcta gtgtagccgt agttagccca ccacttcaag aactctgtag caccgcctac
     5461 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     5521 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     5581 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     5641 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     5701 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     5761 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     5821 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     5881 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     5941 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     6001 cgagtcagtg agcgaggaag cggaaggcga gagtagggaa ctgccaggca tcaaactaag
     6061 cagaaggccc ctgacggatg gcctttttgc gtttctacaa actctttctg tgttgtaaaa
     6121 cgacggccag tcttaagctc gggccccctg ggcggttctg ataacgagta atcgttaatc
     6181 cgcaaataac gtaaaaaccc gcttcggcgg gtttttttat ggggggagtt tagggaaaga
     6241 gcatttgtca gaatatttaa gggcgcctgt cactttgctt gatatatgag aattatttaa
     6301 ccttataaat gagaaaaaag caacgcactt taaataagat acgttgcttt ttcgattgat
     6361 gaacacctat aattaaacta ttcatctatt atttatgatt ttttgtatat acaatatttc
     6421 tagtttgtta aagagaatta agaaaataaa tctcgaaaat aataaaggga aaatcagttt
     6481 ttgatatcaa aattatacat gtcaacgata atacaaaata taatacaaac tataagatgt
     6541 tatcagtatt tattatcatt tagaataaat tttgtgtcgc ccttaattgt gagcggataa
     6601 caattacgag cttcatgcac agtggcgttg actcaatatt ggccattagc catattattc
     6661 attggttata tagcataaat caatattggc tattggccat tgcatacgtt gtatctatat
     6721 cataatatgt acatttatat tggctcatgt ccaatatgac cgccatgtt
//
