LOCUS       sequence-303041-        8329 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 8329)
  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 8329)
  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-SEMA3D-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..8329
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     124..143
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     149..168
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      254..842
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     743..762
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     996..1013
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     1112..1128
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1584..1600
                     /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)."
     enhancer        1767..2146
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2147..2350
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2301..2321
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2347..2371
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        2581..2599
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2581..2598
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     2612..2631
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             5671..5715
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5716..5733
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(5873..5890)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5879..5990
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6108..6128)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6113..6442
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6293..6428
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6355..6374
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6490..6537
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6556..7155
                     /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(6556..6575)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7052..7072
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7285..7406
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7322..7341)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7376..7395
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7459..8250,8251..8319))
                     /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     8082..8101
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8320..8329,1..82))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
       61 tgtatttaga aaaataaaca aataggggtc agtgttacaa ccaattaacc aattctgaac
      121 attatcgcga gcccatttat acctgaatat ggctcataac accccttgct catgaccaaa
      181 atcccttaac gtgagttacg cgcgcgtcgt tccactgagc gtcagacccc gtagaaaaga
      241 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
      301 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
      361 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
      421 tagcccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
      481 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
      541 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
      601 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
      661 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
      721 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
      781 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
      841 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
      901 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag
      961 ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg
     1021 aaggcgagag tagggaactg ccaggcatca aactaagcag aaggcccctg acggatggcc
     1081 tttttgcgtt tctacaaact ctttctgtgt tgtaaaacga cggccagtct taagctcggg
     1141 ccccctgggc ggttctgata acgagtaatc gttaatccgc aaataacgta aaaacccgct
     1201 tcggcgggtt tttttatggg gggagtttag ggaaagagca tttgtcagaa tatttaaggg
     1261 cgcctgtcac tttgcttgat atatgagaat tatttaacct tataaatgag aaaaaagcaa
     1321 cgcactttaa ataagatacg ttgctttttc gattgatgaa cacctataat taaactattc
     1381 atctattatt tatgattttt tgtatataca atatttctag tttgttaaag agaattaaga
     1441 aaataaatct cgaaaataat aaagggaaaa tcagtttttg atatcaaaat tatacatgtc
     1501 aacgataata caaaatataa tacaaactat aagatgttat cagtatttat tatcatttag
     1561 aataaatttt gtgtcgccct taattgtgag cggataacaa ttacgagctt catgcacagt
     1621 ggcgttgact caatattggc cattagccat attattcatt ggttatatag cataaatcaa
     1681 tattggctat tggccattgc atacgttgta tctatatcat aatatgtaca tttatattgg
     1741 ctcatgtcca atatgaccgc catgttggca ttgattattg actagttatt aatagtaatc
     1801 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
     1861 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
     1921 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
     1981 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
     2041 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
     2101 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
     2161 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
     2221 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
     2281 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
     2341 aagcagagct cgtttagtga accgtcagat cgcctggaga cgccatccac gctgttttga
     2401 cctccataga agacaccggg accgatccag cctccgcggc cgggaacggt gcattggaac
     2461 gcggattccc cgtgccaaga gtgacgtaag taccgcctat agagtctata ggcccacccc
     2521 cttggcttcg ttagaacgcg gctacaatta atacataacc ttatgtatca tacacatacg
     2581 atttaggtga cactatagaa taacatccac tttgcctttc tctccacagg tgtccactcc
     2641 caggtccaac tgcacctcgg ttctatcgaa aacgcgcctg ctagcgccac catgaagacc
     2701 atcattgccc tgagctacat cttctgcctg gtgttcgctg cggccgccac actgaagcag
     2761 aacatcccca gactgaagct gacctacaag gacctgctgc tgagcaacag ctgcatccca
     2821 tttctgggca gcagcgaagg cctggacttt cagacactgc tgctggacga ggaacggggc
     2881 agactgctgc ttggagccaa ggaccacatc tttctgctga gcctggtgga cctgaacaag
     2941 aacttcaaga agatctactg gcctgccgcc aaagaacgcg tggaactgtg taaactggcc
     3001 ggcaaggacg ccaataccga gtgcgccaac ttcatcagag tgctgcagcc ctacaacaag
     3061 acccacatct acgtgtgtgg caccggcgcc ttccatccta tctgcggcta tatcgacctg
     3121 ggcgtgtaca aagaggacat catcttcaag ctggacaccc acaacctgga aagcggcagg
     3181 ctgaagtgcc ccttcgatcc tcagcagcct ttcgccagcg tgatgaccga cgagtacctg
     3241 tacagcggaa ccgccagcga cttcctgggc aaagataccg ccttcaccag atctctgggc
     3301 cccacacacg accaccacta catcagaacc gacatcagcg agcactactg gctgaacggc
     3361 gccaagttta tcggcacctt cttcatcccc gacacctaca atcccgacga cgacaagatc
     3421 tatttcttct tccgggaatc ctctcaagag ggcagcacca gcgacaagac catcctgagc
     3481 agagtgggca gagtgtgcaa gaacgatgtc ggcggacagc ggagcctgat caacaagtgg
     3541 accaccttcc tgaaggcccg gctgatctgt agcatccctg gatctgatgg cgccgatacc
     3601 tacttcgacg agctgcagga tatctacctg ctgcctacca gggacgagag aaaccctgtg
     3661 gtgtacggcg tgttcaccac caccagctcc atctttaagg gcagcgccgt gtgcgtgtac
     3721 agcatggccg atatcagagc cgtgttcaac ggcccctacg ctcacaaaga gtccgccgat
     3781 catagatggg tgcagtacga cggcagaatc ccctatccta gacctggcac atgccccagc
     3841 aagacctacg atcccctgat caagagcacc cgggacttcc ccgatgacgt gatcagcttc
     3901 atcaagcggc actccgtgat gtacaagagc gtgtaccctg tggctggcgg ccctaccttc
     3961 aagagaatca acgtggacta ccggctgacc cagatcgtgg tggatcatgt gatcgccgag
     4021 gacggccagt acgatgtgat gttcctggga acagacatcg gcaccgtgct gaaggtggtg
     4081 tccatcagca aagaaaagtg gaacatggaa gaggtggtgc tcgaggaact gcagatcttt
     4141 aagcacagca gcatcatcct caacatggaa ctgagcctga agcagcagca gctgtacatc
     4201 ggctccagag atggcctggt gcagctgtcc ctgcacagat gtgataccta cggcaaggcc
     4261 tgcgccgatt gctgccttgc cagagatcct tactgtgcct gggatggcaa cgcctgcagc
     4321 agatacgccc caacatctaa gagaagggcc agacggcagg acgtgaagta cggcgatcct
     4381 atcacacagt gctgggacat cgaggacagc atcagccacg agacagccga cgagaaagtg
     4441 atcttcggca tcgagttcaa cagcaccttc ctcgagtgca tccccaagag ccagcaggcc
     4501 accatcaagt ggtacatcca gagaagcggc gacgagcaca gagaggaact gaagccagac
     4561 gagcggatca tcaagaccga gtacggcctg ctgatcagaa gcctgcagaa aaaggacagc
     4621 ggcatgtact actgcaaggc ccaagagcac accttcatcc acaccatcgt gaaactgacc
     4681 ctgaacgtga tcgagaacga gcagatggaa aacacccagc gggccgagca cgaagagggc
     4741 aaagttaagg atctgctcgc cgagagccgg ctgcggtaca aggactatat ccagatcctg
     4801 tctagcccca atttcagcct ggaccagtac tgcgaacaga tgtggcacag agaaaagcgg
     4861 cggcagagaa acaaaggcgg cccaaagtgg aagcacatgc aagagatgaa gaaaaagcgg
     4921 aacagacggc accaccgcga cctggatgaa ctgcctagag ctgtggctac aggcgcgccg
     4981 ggcggcgaca agacccacac ctgccccccc tgcccagctc cagaactgct gggagggcct
     5041 tccgtgttcc tgttcccccc caagcccaag gacaccctga tgatcagcag gacccccgag
     5101 gtgacctgcg tggtggtggc cgtgtcccac gaggacccag aggtgaagtt caactggtac
     5161 gtggacggcg tggaggtgca caacgccaag accaagccca gagaggagca gtacaacagc
     5221 acctacaggg tggtgtccgt gctgaccgtg ctgcaccagg actggctgaa cggcaaagaa
     5281 tacaagtgca aagtctccaa caaggccctg gctgccccaa tcgaaaagac aatcagcaag
     5341 gccaagggcc agccacggga gccccaggtg tacaccctgc cccccagccg ggaggagatg
     5401 accaagaacc aggtgtccct gacctgtctg gtgaagggct tctaccccag cgatatcgcc
     5461 gtggagtggg agagcaacgg ccagcccgag aacaactaca agaccacccc cccagtgctg
     5521 gacagcgacg gcagcttctt cctgtacagc aagctgaccg tggacaagtc caggtggcag
     5581 cagggcaacg tgttcagctg cagcgtgatg cacgaggccc tgcacaacca ctacacccag
     5641 aagtccctga gcctgagccc cggcggcagc ggtcttaatg atattttcga ggctcagaag
     5701 attgagtggc atgaacacca ccaccatcac cactaaggcc ggcctgtagt cactggcaca
     5761 acgcgttacc cggtaagcca atcgggtata cacggtcgtc atactgcaga cagggttctt
     5821 ctactttgca agatagtctt gagtagtaaa ataaatagat agagaaaatc agcctcgact
     5881 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg
     5941 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc acaacactca
     6001 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     6061 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
     6121 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     6181 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     6241 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     6301 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     6361 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
     6421 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
     6481 tcagcacgtg ttgacaatta atcatcggca tagtatatcg gcatagtata atacgacaag
     6541 gtgaggaact aaaccatgac cgagtacaag cccacggtgc gcctcgccac ccgcgacgac
     6601 gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc cacgcgccac
     6661 accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg
     6721 cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc
     6781 tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg
     6841 gccgagttga gcggttcccg gctggccgcg cagcaacaga tggaaggcct cctggcgccg
     6901 caccggccca aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag
     6961 ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg
     7021 gtgcccgcct tcctggagac atccgcgccc cgcaacctcc ccttctacga gcggctcggc
     7081 ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc
     7141 aagcccggtg cctgacacgt gctacgagat ttcgattcca ccgccgcctt ctatgaaagg
     7201 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     7261 atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa
     7321 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     7381 ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc
     7441 ttggcgtaat catggtcatt accaatgctt aatcagtgag gcacctatct cagcgatctg
     7501 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     7561 gggcttacca tctggcccca gcgctgcgat gataccgcga gaaccacgct caccggctcc
     7621 ggatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     7681 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     7741 agttaatagt ttgcgcaacg ttgttgccat cgctacaggc atcgtggtgt cacgctcgtc
     7801 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     7861 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     7921 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     7981 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     8041 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     8101 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     8161 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     8221 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     8281 aaagggaata agggcgacac ggaaatgttg aatactcata ttcttcctt
//
