LOCUS       sequence-430452-        6757 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 6757)
  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 6757)
  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-KIR2DL5B-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6757
                     /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             2333..2377
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             2378..2395
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2535..2552)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2541..2652
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2770..2790)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2775..3104
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2955..3090
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3017..3036
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3152..3199
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3218..3817
                     /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(3218..3237)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3714..3734
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    3947..4068
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3984..4003)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4038..4057
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(4121..4912,4913..4981))
                     /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     4744..4763
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4982..5073)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5115..5134
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5140..5159
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      5245..5833
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5734..5753
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5987..6004
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     6102..6119
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6103..6119
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6575..6591
                     /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 cccatgaagg tggacaagat aagcctctgc tgagcgcctg
     1021 gccttctgct gttgttccta gaggcggaca cgtgaccctg ctgtgtagaa gcagactggg
     1081 cttcaccatc ttcagcctgt acaaagagga cggcgtcccc gttcctgagc tgtacaacaa
     1141 gatcttctgg aagtccatcc tgatgggccc cgtgacacca gctcacgccg gcacatatag
     1201 atgcagaggc tctcacccta gaagccccat tgagtggtcc gctcctagca accctctggt
     1261 catcgtggtc accggcctgt ttggcaagcc ttctctgtct gctcagcctg gacctacagt
     1321 gcggacaggc gagaatgtga ccctgagctg cagcagcaga tccagcttcg acatgtacca
     1381 cctgagcaga gagggcagag cccacgaacc tagactgcct gctgtgcctt ccgtgaatgg
     1441 caccttccag gccgattttc cactgggacc tgccacacac ggcggcacct acacatgttt
     1501 tggcagcctg cacgacagcc cctacgagtg gtctgatcct tctgatccac tgctggtgtc
     1561 cgtgaccggc aatagcagta gcagcagctc tagccctacc gagcctagca gcaagaccgg
     1621 catcagaagg catggcgcgc cgggcggcga caagacccac acctgccccc cctgcccagc
     1681 tccagaactg ctgggagggc cttccgtgtt cctgttcccc cccaagccca aggacaccct
     1741 gatgatcagc aggacccccg aggtgacctg cgtggtggtg gccgtgtccc acgaggaccc
     1801 agaggtgaag ttcaactggt acgtggacgg cgtggaggtg cacaacgcca agaccaagcc
     1861 cagagaggag cagtacaaca gcacctacag ggtggtgtcc gtgctgaccg tgctgcacca
     1921 ggactggctg aacggcaaag aatacaagtg caaagtctcc aacaaggccc tggctgcccc
     1981 aatcgaaaag acaatcagca aggccaaggg ccagccacgg gagccccagg tgtacaccct
     2041 gccccccagc cgggaggaga tgaccaagaa ccaggtgtcc ctgacctgtc tggtgaaggg
     2101 cttctacccc agcgatatcg ccgtggagtg ggagagcaac ggccagcccg agaacaacta
     2161 caagaccacc cccccagtgc tggacagcga cggcagcttc ttcctgtaca gcaagctgac
     2221 cgtggacaag tccaggtggc agcagggcaa cgtgttcagc tgcagcgtga tgcacgaggc
     2281 cctgcacaac cactacaccc agaagtccct gagcctgagc cccggcggca gcggtcttaa
     2341 tgatattttc gaggctcaga agattgagtg gcatgaacac caccaccatc accactaagg
     2401 ccggcctgta gtcactggca caacgcgtta cccggtaagc caatcgggta tacacggtcg
     2461 tcatactgca gacagggttc ttctactttg caagatagtc ttgagtagta aaataaatag
     2521 atagagaaaa tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
     2581 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     2641 ggaaattgca tcacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     2701 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     2761 taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     2821 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     2881 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     2941 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     3001 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc
     3061 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt
     3121 gtatatccat tttcggatct gatcagcacg tgttgacaat taatcatcgg catagtatat
     3181 cggcatagta taatacgaca aggtgaggaa ctaaaccatg accgagtaca agcccacggt
     3241 gcgcctcgcc acccgcgacg acgtccccag ggccgtacgc accctcgccg ccgcgttcgc
     3301 cgactacccc gccacgcgcc acaccgtcga tccggaccgc cacatcgagc gggtcaccga
     3361 gctgcaagaa ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga
     3421 cgacggcgcc gcggtggcgg tctggaccac gccggagagc gtcgaagcgg gggcggtgtt
     3481 cgccgagatc ggcccgcgca tggccgagtt gagcggttcc cggctggccg cgcagcaaca
     3541 gatggaaggc ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc tggccaccgt
     3601 cggcgtctcg cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc tccccggagt
     3661 ggaggcggcc gagcgcgccg gggtgcccgc cttcctggag acatccgcgc cccgcaacct
     3721 ccccttctac gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc
     3781 gcgcacctgg tgcatgaccc gcaagcccgg tgcctgacac gtgctacgag atttcgattc
     3841 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     3901 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc
     3961 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     4021 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat
     4081 accgtcgacc tctagctaga gcttggcgta atcatggtca ttaccaatgc ttaatcagtg
     4141 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     4201 tgtagataac tacgatacgg gagggcttac catctggccc cagcgctgcg atgataccgc
     4261 gagaaccacg ctcaccggct ccggatttat cagcaataaa ccagccagcc ggaagggccg
     4321 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     4381 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc atcgctacag
     4441 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     4501 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     4561 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     4621 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     4681 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     4741 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     4801 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     4861 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     4921 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     4981 tattcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     5041 acatatttga atgtatttag aaaaataaac aaataggggt cagtgttaca accaattaac
     5101 caattctgaa cattatcgcg agcccattta tacctgaata tggctcataa caccccttgc
     5161 tcatgaccaa aatcccttaa cgtgagttac gcgcgcgtcg ttccactgag cgtcagaccc
     5221 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     5281 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     5341 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     5401 gtagccgtag ttagcccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     5461 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     5521 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     5581 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     5641 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     5701 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     5761 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     5821 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     5881 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     5941 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     6001 cgaggaagcg gaaggcgaga gtagggaact gccaggcatc aaactaagca gaaggcccct
     6061 gacggatggc ctttttgcgt ttctacaaac tctttctgtg ttgtaaaacg acggccagtc
     6121 ttaagctcgg gccccctggg cggttctgat aacgagtaat cgttaatccg caaataacgt
     6181 aaaaacccgc ttcggcgggt ttttttatgg ggggagttta gggaaagagc atttgtcaga
     6241 atatttaagg gcgcctgtca ctttgcttga tatatgagaa ttatttaacc ttataaatga
     6301 gaaaaaagca acgcacttta aataagatac gttgcttttt cgattgatga acacctataa
     6361 ttaaactatt catctattat ttatgatttt ttgtatatac aatatttcta gtttgttaaa
     6421 gagaattaag aaaataaatc tcgaaaataa taaagggaaa atcagttttt gatatcaaaa
     6481 ttatacatgt caacgataat acaaaatata atacaaacta taagatgtta tcagtattta
     6541 ttatcattta gaataaattt tgtgtcgccc ttaattgtga gcggataaca attacgagct
     6601 tcatgcacag tggcgttgac tcaatattgg ccattagcca tattattcat tggttatata
     6661 gcataaatca atattggcta ttggccattg catacgttgt atctatatca taatatgtac
     6721 atttatattg gctcatgtcc aatatgaccg ccatgtt
//
