LOCUS       sequence-431376-        6994 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 6994)
  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 6994)
  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-KIR3DL3.1-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6994
                     /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             2570..2614
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             2615..2632
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2772..2789)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2778..2889
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3007..3027)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3012..3341
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3192..3327
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3254..3273
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3389..3436
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3455..4054
                     /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(3455..3474)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3951..3971
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4184..4305
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4221..4240)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4275..4294
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(4358..5149,5150..5218))
                     /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     4981..5000
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5219..5310)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5352..5371
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5377..5396
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      5482..6070
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5971..5990
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6224..6241
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     6339..6356
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6340..6356
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6812..6828
                     /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 cccaagataa gccttttctg tctgcttggc ctggcaccgt
     1021 ggtgtctgaa ggacagcatg tgaccctgca gtgcagaagc agactgggct tcaacgagtt
     1081 cagcctgagc aaagaggacg gcatgcctgt gcctgagctg tacaaccgga tcttccggaa
     1141 cagcttcctg atgggccctg tgacaccagc tcacgccggc acatacagat gctgtagctc
     1201 tcaccctcac agccccacag gatggtccgc tccatctaac cctgtggtca tcatggtcac
     1261 cggcgtgcac agaaagcctt ctctgcttgc ccatcctgga cctctggtca agtctggcga
     1321 gacagtgatt ctgcagtgtt ggagcgacgt ccgcttcgag agattcctgc tgcacagaga
     1381 gggcatcaca gaggaccctc tgagactcgt gggacagctg catgatgccg gcagccaagt
     1441 gaattacagc atgggcccaa tgacacccgc actggccggc acctatagat gtttcggcag
     1501 cgtgacccat ctgccttacg agctgtctgc ccctagcgac cctctggata ttgtggttgt
     1561 gggcctgtac ggcaagccta gcctttctgc tcagcctgga cctacagtgc aggccggcga
     1621 aaatgtgaca ctgagctgtt ccagcagaag cctgttcgac atctaccacc tgtccagaga
     1681 agccgaagcc ggcgaactga gactgacagc cgtgctgaga gtgaacggca ccttccaggc
     1741 caattttcca ctgggaccag tgacacacgg cggcaattac cggtgcttcg gctcttttag
     1801 agccctgcct catgcttgga gcgatccctc tgatcctctg cctgtgtctg tgaccggcaa
     1861 cagcagaaat ggcgcgccgg gcggcgacaa gacccacacc tgccccccct gcccagctcc
     1921 agaactgctg ggagggcctt ccgtgttcct gttccccccc aagcccaagg acaccctgat
     1981 gatcagcagg acccccgagg tgacctgcgt ggtggtggcc gtgtcccacg aggacccaga
     2041 ggtgaagttc aactggtacg tggacggcgt ggaggtgcac aacgccaaga ccaagcccag
     2101 agaggagcag tacaacagca cctacagggt ggtgtccgtg ctgaccgtgc tgcaccagga
     2161 ctggctgaac ggcaaagaat acaagtgcaa agtctccaac aaggccctgg ctgccccaat
     2221 cgaaaagaca atcagcaagg ccaagggcca gccacgggag ccccaggtgt acaccctgcc
     2281 ccccagccgg gaggagatga ccaagaacca ggtgtccctg acctgtctgg tgaagggctt
     2341 ctaccccagc gatatcgccg tggagtggga gagcaacggc cagcccgaga acaactacaa
     2401 gaccaccccc ccagtgctgg acagcgacgg cagcttcttc ctgtacagca agctgaccgt
     2461 ggacaagtcc aggtggcagc agggcaacgt gttcagctgc agcgtgatgc acgaggccct
     2521 gcacaaccac tacacccaga agtccctgag cctgagcccc ggcggcagcg gtcttaatga
     2581 tattttcgag gctcagaaga ttgagtggca tgaacaccac caccatcacc actaaggccg
     2641 gcctgtagtc actggcacaa cgcgttaccc ggtaagccaa tcgggtatac acggtcgtca
     2701 tactgcagac agggttcttc tactttgcaa gatagtcttg agtagtaaaa taaatagata
     2761 gagaaaatca gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc
     2821 cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga
     2881 aattgcatca caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     2941 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa
     3001 ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     3061 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     3121 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     3181 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     3241 gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga
     3301 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta
     3361 tatccatttt cggatctgat cagcacgtgt tgacaattaa tcatcggcat agtatatcgg
     3421 catagtataa tacgacaagg tgaggaacta aaccatgacc gagtacaagc ccacggtgcg
     3481 cctcgccacc cgcgacgacg tccccagggc cgtacgcacc ctcgccgccg cgttcgccga
     3541 ctaccccgcc acgcgccaca ccgtcgatcc ggaccgccac atcgagcggg tcaccgagct
     3601 gcaagaactc ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga
     3661 cggcgccgcg gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc
     3721 cgagatcggc ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc agcaacagat
     3781 ggaaggcctc ctggcgccgc accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg
     3841 cgtctcgccc gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga
     3901 ggcggccgag cgcgccgggg tgcccgcctt cctggagaca tccgcgcccc gcaacctccc
     3961 cttctacgag cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg
     4021 cacctggtgc atgacccgca agcccggtgc ctgacacgtg ctacgagatt tcgattccac
     4081 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
     4141 cctccagcgc ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc
     4201 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     4261 actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc
     4321 gtcgacctct agctagagct tggcgtaatc atggtcatta ccaatgctta atcagtgagg
     4381 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     4441 agataactac gatacgggag ggcttaccat ctggccccag cgctgcgatg ataccgcgag
     4501 aaccacgctc accggctccg gatttatcag caataaacca gccagccgga agggccgagc
     4561 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     4621 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatc gctacaggca
     4681 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     4741 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     4801 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     4861 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     4921 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     4981 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     5041 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     5101 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     5161 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatat
     5221 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     5281 tatttgaatg tatttagaaa aataaacaaa taggggtcag tgttacaacc aattaaccaa
     5341 ttctgaacat tatcgcgagc ccatttatac ctgaatatgg ctcataacac cccttgctca
     5401 tgaccaaaat cccttaacgt gagttacgcg cgcgtcgttc cactgagcgt cagaccccgt
     5461 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     5521 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     5581 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
     5641 gccgtagtta gcccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     5701 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     5761 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     5821 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     5881 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     5941 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     6001 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     6061 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     6121 tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt
     6181 tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga
     6241 ggaagcggaa ggcgagagta gggaactgcc aggcatcaaa ctaagcagaa ggcccctgac
     6301 ggatggcctt tttgcgtttc tacaaactct ttctgtgttg taaaacgacg gccagtctta
     6361 agctcgggcc ccctgggcgg ttctgataac gagtaatcgt taatccgcaa ataacgtaaa
     6421 aacccgcttc ggcgggtttt tttatggggg gagtttaggg aaagagcatt tgtcagaata
     6481 tttaagggcg cctgtcactt tgcttgatat atgagaatta tttaacctta taaatgagaa
     6541 aaaagcaacg cactttaaat aagatacgtt gctttttcga ttgatgaaca cctataatta
     6601 aactattcat ctattattta tgattttttg tatatacaat atttctagtt tgttaaagag
     6661 aattaagaaa ataaatctcg aaaataataa agggaaaatc agtttttgat atcaaaatta
     6721 tacatgtcaa cgataataca aaatataata caaactataa gatgttatca gtatttatta
     6781 tcatttagaa taaattttgt gtcgccctta attgtgagcg gataacaatt acgagcttca
     6841 tgcacagtgg cgttgactca atattggcca ttagccatat tattcattgg ttatatagca
     6901 taaatcaata ttggctattg gccattgcat acgttgtatc tatatcataa tatgtacatt
     6961 tatattggct catgtccaat atgaccgcca tgtt
//
