LOCUS       sequence-424406-        8407 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of secreted protein fused to 
            Fc(DAPA)-Avi-6xHis..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8407)
  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 8407)
  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-SEMA3F-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..8407
                     /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"
     RBS             3100..3108
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     CDS             3983..4027
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             4028..4045
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4185..4202)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4191..4302
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4420..4440)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4425..4754
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4605..4740
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4667..4686
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4802..4849
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4868..5467
                     /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(4868..4887)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5364..5384
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5597..5718
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5634..5653)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5688..5707
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(5771..6562,6563..6631))
                     /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     6394..6413
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6632..6723)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6765..6784
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6790..6809
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      6895..7483
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7384..7403
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7637..7654
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     7752..7769
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7753..7769
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    8225..8241
                     /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 ccagctttcc cacacaagat catctgcctg ccacacctag
     1021 agtgcggctg agctttaaag agctgaaggc caccggcaca gcccactttt tcaactttct
     1081 gctgaacacc accgactacc gcatcctgct gaaggacgag gaccacgatc gtatgtacgt
     1141 gggcagcaag gactacgtgc tgagcctgga cctgcacgac atcaacagag agcccctgat
     1201 catccattgg gccgcctctc cacagagaat cgaggaatgt gtgctgagcg gcaaggacgt
     1261 gaacggcgag tgtggaaact tcgtcagact gatccagcct tggaacagaa cccacctgta
     1321 cgtgtgtggc accggcgcct acaatcctat gtgcacctac gtgaacagag gccgcagagc
     1381 acaggctacc ccttggacac agacacaggc cgttagagga agaggctcca gagctacaga
     1441 tggcgctctg aggcctatgc ctaccgctcc tagacaggac tacatcttct acctggaacc
     1501 tgagcggctg gaatccggca agggcaagtg tccttacgac cccaagctgg atacagccag
     1561 cgctctgatc aacgaggaac tgtatgccgg cgtgtacatc gacttcatgg gcaccgatgc
     1621 cgccatcttc agaaccctgg gaaagcagac cgccatgcgg accgaccagt acaacagcag
     1681 atggctgaac gaccccagct tcatccacgc cgagctgatc cctgatagcg ccgagagaaa
     1741 cgacgacaag ctgtacttct tcttccgcga acgctctgcc gaggctcctc agtctcctgc
     1801 cgtgtatgcc agaatcggca gaatctgcct gaacgacgat ggcggccact gctgtctggt
     1861 caacaagtgg tctaccttcc tgaaggcccg gctcgtttgt tctgtgcctg gcgaggatgg
     1921 catcgagaca cacttcgatg agctgcagga cgtgttcgtg cagcagaccc aggatgtgcg
     1981 gaatcctgtg atctacgccg tgtttacaag cagcggcagc gtgttcagag gcagcgccgt
     2041 gtgtgtgtac tccatggccg acatcagaat ggtgttcaac ggccccttcg ctcacaaaga
     2101 gggccccaat taccagtgga tgcccttcag cggcaagatg ccctatccta gacctggcac
     2161 atgtcctggc ggcaccttca cacctagcat gaagtccacc aaggattacc ccgacgaagt
     2221 gatcaacttc atgcggagcc atcctctgat gtaccaggcc gtgtatcccc tgcagagaag
     2281 gcctctggtt gttagaacag gcgcccctta cagactgacc acaatcgctg tggatcaggt
     2341 ggacgccgcc gatggaagat acgaggtgct gttcctgggc acagacagag gcaccgtgca
     2401 gaaagtgatc gtgctgccca aggacgacca agagctggaa gaactgatgc tggaagaggt
     2461 ggaagtgttc aaggaccccg ctcctgtgaa aaccatgacc atcagcagca agcggcagca
     2521 gctgtatgtg gcttctgccg tgggagtgac acacctgtct ctgcacagat gccaggctta
     2581 tggcgccgct tgtgccgatt gctgtctcgc cagagatcct tattgtgcct gggatggcca
     2641 ggcctgtagc agatacacag ccagctccaa gagaagatcc cgcagacagg acgtgcggca
     2701 cggcaatcct atcagacagt gcagaggctt caacagcaac gccaacaaga acgccgtgga
     2761 aagcgtgcag tatggcgtgg ccggatctgc cgcctttctt gagtgtcagc ctagaagccc
     2821 tcaggccacc gtgaagtggc tgtttcagag agatcccggc gaccggcgga gagagattag
     2881 agccgaggat agattcctga gaaccgagca gggactgctg ctgagagcac tgcaactgag
     2941 cgatagaggc ctgtactctt gcaccgccac cgagaacaac ttcaagcacg tggtcaccag
     3001 agtccagctg cacgtgctgg gaagagatgc tgtgcatgcc gctctgttcc ctccactgtc
     3061 tatgtctgct cctcctccac ctggtgctgg acctccaaca cctccttatc aagaactggc
     3121 ccagctgctg gctcagcctg aagtgggact gattcaccag tactgccaag gctactggcg
     3181 ccacgttcca ccttcaccta gagaagctcc aggcgctcca agaagccctg agcctcagga
     3241 tcagaagaag cccagaaaca gacggcacca tcctccagac acaggcgcgc cgggcggcga
     3301 caagacccac acctgccccc cctgcccagc tccagaactg ctgggagggc cttccgtgtt
     3361 cctgttcccc cccaagccca aggacaccct gatgatcagc aggacccccg aggtgacctg
     3421 cgtggtggtg gccgtgtccc acgaggaccc agaggtgaag ttcaactggt acgtggacgg
     3481 cgtggaggtg cacaacgcca agaccaagcc cagagaggag cagtacaaca gcacctacag
     3541 ggtggtgtcc gtgctgaccg tgctgcacca ggactggctg aacggcaaag aatacaagtg
     3601 caaagtctcc aacaaggccc tggctgcccc aatcgaaaag acaatcagca aggccaaggg
     3661 ccagccacgg gagccccagg tgtacaccct gccccccagc cgggaggaga tgaccaagaa
     3721 ccaggtgtcc ctgacctgtc tggtgaaggg cttctacccc agcgatatcg ccgtggagtg
     3781 ggagagcaac ggccagcccg agaacaacta caagaccacc cccccagtgc tggacagcga
     3841 cggcagcttc ttcctgtaca gcaagctgac cgtggacaag tccaggtggc agcagggcaa
     3901 cgtgttcagc tgcagcgtga tgcacgaggc cctgcacaac cactacaccc agaagtccct
     3961 gagcctgagc cccggcggca gcggtcttaa tgatattttc gaggctcaga agattgagtg
     4021 gcatgaacac caccaccatc accactaagg ccggcctgta gtcactggca caacgcgtta
     4081 cccggtaagc caatcgggta tacacggtcg tcatactgca gacagggttc ttctactttg
     4141 caagatagtc ttgagtagta aaataaatag atagagaaaa tcagcctcga ctgtgccttc
     4201 tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc
     4261 cactcccact gtcctttcct aataaaatga ggaaattgca tcacaacact caaccctatc
     4321 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     4381 gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt
     4441 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     4501 cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     4561 atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc
     4621 cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg
     4681 ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
     4741 taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct gatcagcacg
     4801 tgttgacaat taatcatcgg catagtatat cggcatagta taatacgaca aggtgaggaa
     4861 ctaaaccatg accgagtaca agcccacggt gcgcctcgcc acccgcgacg acgtccccag
     4921 ggccgtacgc accctcgccg ccgcgttcgc cgactacccc gccacgcgcc acaccgtcga
     4981 tccggaccgc cacatcgagc gggtcaccga gctgcaagaa ctcttcctca cgcgcgtcgg
     5041 gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg tctggaccac
     5101 gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca tggccgagtt
     5161 gagcggttcc cggctggccg cgcagcaaca gatggaaggc ctcctggcgc cgcaccggcc
     5221 caaggagccc gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc agggcaaggg
     5281 tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg gggtgcccgc
     5341 cttcctggag acatccgcgc cccgcaacct ccccttctac gagcggctcg gcttcaccgt
     5401 caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc gcaagcccgg
     5461 tgcctgacac gtgctacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
     5521 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     5581 gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     5641 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     5701 actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta
     5761 atcatggtca ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     5821 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     5881 catctggccc cagcgctgcg atgataccgc gagaaccacg ctcaccggct ccggatttat
     5941 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     6001 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     6061 gtttgcgcaa cgttgttgcc atcgctacag gcatcgtggt gtcacgctcg tcgtttggta
     6121 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     6181 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     6241 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     6301 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     6361 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     6421 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     6481 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     6541 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     6601 taagggcgac acggaaatgt tgaatactca tattcttcct ttttcaatat tattgaagca
     6661 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     6721 aaataggggt cagtgttaca accaattaac caattctgaa cattatcgcg agcccattta
     6781 tacctgaata tggctcataa caccccttgc tcatgaccaa aatcccttaa cgtgagttac
     6841 gcgcgcgtcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     6901 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     6961 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     7021 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttagcccacc acttcaagaa
     7081 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     7141 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     7201 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     7261 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     7321 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     7381 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     7441 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     7501 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     7561 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     7621 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaaggcgaga gtagggaact
     7681 gccaggcatc aaactaagca gaaggcccct gacggatggc ctttttgcgt ttctacaaac
     7741 tctttctgtg ttgtaaaacg acggccagtc ttaagctcgg gccccctggg cggttctgat
     7801 aacgagtaat cgttaatccg caaataacgt aaaaacccgc ttcggcgggt ttttttatgg
     7861 ggggagttta gggaaagagc atttgtcaga atatttaagg gcgcctgtca ctttgcttga
     7921 tatatgagaa ttatttaacc ttataaatga gaaaaaagca acgcacttta aataagatac
     7981 gttgcttttt cgattgatga acacctataa ttaaactatt catctattat ttatgatttt
     8041 ttgtatatac aatatttcta gtttgttaaa gagaattaag aaaataaatc tcgaaaataa
     8101 taaagggaaa atcagttttt gatatcaaaa ttatacatgt caacgataat acaaaatata
     8161 atacaaacta taagatgtta tcagtattta ttatcattta gaataaattt tgtgtcgccc
     8221 ttaattgtga gcggataaca attacgagct tcatgcacag tggcgttgac tcaatattgg
     8281 ccattagcca tattattcat tggttatata gcataaatca atattggcta ttggccattg
     8341 catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc aatatgaccg
     8401 ccatgtt
//
