LOCUS       sequence-303161-        9259 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 9259)
  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 9259)
  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-CNTN5-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..9259
                     /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     1111..1128
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     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             6601..6645
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6646..6663
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6803..6820)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6809..6920
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(7038..7058)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        7043..7372
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7223..7358
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7285..7304
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7420..7467
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7486..8085
                     /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(7486..7505)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7982..8002
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    8215..8336
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8252..8271)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8306..8325
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(8389..9180,9181..9249))
                     /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     9012..9031
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(9250..9259,1..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 cggccgccga gtattctaag
     2761 tctctgcctg gcctgagcac cagctatgcc gctctgctga gaatcaagaa gtccagcagc
     2821 agctccctgt tcggcagcaa gaccagacct agatacagca gccctagcct gggcaccctg
     2881 tctgcctctt ctccatcttg gcttggcgcc gctcagaact actacagccc catcaacctg
     2941 taccacagca gcgacgcctt caagcaggat gagtccgtgg attacggccc cgtgttcgtg
     3001 caagagcccg acgacatcat cttccccacc gacagcgacg agaagaaggt ggccctgaat
     3061 tgcgaagtgc ggggcaatcc tgtgcctagc taccggtggc tgagaaacgg cacagagatc
     3121 gacctggaaa gcgactaccg gtacagcctg atcgacggca ccttcatcat cagcaacccc
     3181 agcgaggcca aggacagcgg ccattatcag tgcctggcca ccaataccgt gggcagcatt
     3241 ctgagcagag aggccacact gcagttcgcc tacctgggca atttcagcgg cagaaccaga
     3301 tccgccgtgt ccgtcagaga aggacaaggc gtggtgctga tgtgcagccc tccacctcac
     3361 agccccgaga tcatctacag ctgggtgttc aacgagttcc ccagcttcgt ggccgaggac
     3421 agcagacggt tcatcagcca agagacaggc aatctgtaca tcagcaaggt gcagaccagc
     3481 gacgtgggct cctacatctg cctggtcaag aacaccgtga ccaacgccag agtgctgagc
     3541 cctcctacac ctctgacact gagaaatgac ggcgtgatgg gcgagtacga gcccaagatc
     3601 gaggtgcact tcccctttac tgtgaccgcc gccaagggca ccaccgtgaa gatggaatgt
     3661 ttcgccctgg gcaaccccgt gcctaccatc acatggatga aggtcaacgg ctacatcccc
     3721 agcaaggccc gcctgagaaa atctcaggcc gtgctggaaa tccccaacgt gcagctggat
     3781 gacgccggca tctatgagtg cagagccgag aactcccggg gcaagaatag cttcagaggc
     3841 cagctgcagg tctacacata ccctcactgg gtcgagaagc tgaacgatac ccagctggat
     3901 agcggcagcc ctctgagatg ggagtgtaaa gccaccggca agcccagacc aacctacaga
     3961 tggctgaaga acggcgtgcc actgagccca cagagcagag tggaaatggt caatggcgtc
     4021 ctgatgatcc acaacgtgaa ccagagcgac gccggaatgt accagtgtct ggccgagaac
     4081 aaatacggcg ccatctacgc ctctgccgag ctgaagattc tggccagcgc tcccacattc
     4141 gccctgaacc agctgaagaa aaccatcatc gtgaccaagg accaagaggt ggtcatcgag
     4201 tgcaagcctc agggcagccc taagccaacc atcagctgga agaaaggcga cagagccgtg
     4261 cgcgagaaca agcggattgc catcctgcct gatggcagcc tgagaatcct gaacgccagc
     4321 aagagcgacg agggcaaata cgtgtgcagg ggcgagaatg tgtttggcag cgccgagatt
     4381 atcgccagcc tgagcgtgaa agagcccacc agaatcgagc tgacccctaa gagaacagag
     4441 ctgaccgtgg gcgagagcat cgtgctgaat tgcaaggcca tccacgatgc cagcctggac
     4501 gtgacctttt actggaccct gaagggccag cctatcgact tcgaggaaga aggcggacac
     4561 ttcgagtcca tcagagccca ggcctctagc gccgatctga tgatcagaaa tatcctgctg
     4621 atgcacgccg gaagatacgg ctgcagagtg cagacaacag ccgacagcgt gtccgatgaa
     4681 gccgagctgc ttgttagagg ccctcctgga ccaccaggca tcgtgatcgt ggaagagatc
     4741 accgagagca ccgccacact gtcttggagc cctgccgccg ataatcacag ccctatcagc
     4801 agctacaacc tgcaggccag atctcccttt agcctcggct ggcagaccgt gaaaaccgtg
     4861 cctgagatca tcaccggcga catggaatct gccatggccg tggacctgaa tccttgggtc
     4921 gagtatgagt tcagagtggt ggccacaaat cccatcggca ccggcgatcc tagcacaccc
     4981 agcagaatga tccggaccaa tgaggccgtg cctaagacag cccctaccaa cgtgtccggc
     5041 agatctggca gaaggcacga gctggttatt gcctgggagc ccgtgtccga ggaattccag
     5101 aatggcgaag gctttggcta catcgtggcc ttcagaccca atggcaccag aggctggaaa
     5161 gaaaagatgg tcaccagctc cgaggccagc aagttcatct accgggatga gagcgtgcca
     5221 ccactgacac ccttcgaagt gaaagtgggc gtgtacaaca acaaaggcga cggccctttc
     5281 agccagatcg tggtcatttg ttctgccgag ggcgagcctt ctgctgcccc aacagatgtg
     5341 aaagccacct ccgtgtctgt gtccgagatc ctggtggcct ggaagcacat caaagagtcc
     5401 ctgggcagac ctcagggctt cgaagtcggc tactggaagg acatggaaca agaggacacc
     5461 gccgagacag tgaaaacaag aggcaacgag agcttcgtga tcctgaccgg actggaaggc
     5521 aacacactgt accactttac cgtgcgggcc tacaacggcg ctggatatgg acctcctagc
     5581 tctgaggtgt ccgccaccac caagaagtct ccaccatctc aggctccctc caatctcaga
     5641 tgggagcagc agggatctca ggtgtccctc ggatgggagc ctgtgatccc tctggccaat
     5701 gagtctgagg tcgtgggcta caaggtgttc taccggcaag agggccacag caacagccaa
     5761 gtgatcgaga cacagaaact gcaggccgtg gtgccacttc ctgatgccgg ggtgtacatc
     5821 atcgaagtcc gggcctattc tgaaggcgga gatggcacag ccagcagcca gattagagtg
     5881 ccatcttaca gcggcggcaa aggcgcgccg ggcggcgaca agacccacac ctgccccccc
     5941 tgcccagctc cagaactgct gggagggcct tccgtgttcc tgttcccccc caagcccaag
     6001 gacaccctga tgatcagcag gacccccgag gtgacctgcg tggtggtggc cgtgtcccac
     6061 gaggacccag aggtgaagtt caactggtac gtggacggcg tggaggtgca caacgccaag
     6121 accaagccca gagaggagca gtacaacagc acctacaggg tggtgtccgt gctgaccgtg
     6181 ctgcaccagg actggctgaa cggcaaagaa tacaagtgca aagtctccaa caaggccctg
     6241 gctgccccaa tcgaaaagac aatcagcaag gccaagggcc agccacggga gccccaggtg
     6301 tacaccctgc cccccagccg ggaggagatg accaagaacc aggtgtccct gacctgtctg
     6361 gtgaagggct tctaccccag cgatatcgcc gtggagtggg agagcaacgg ccagcccgag
     6421 aacaactaca agaccacccc cccagtgctg gacagcgacg gcagcttctt cctgtacagc
     6481 aagctgaccg tggacaagtc caggtggcag cagggcaacg tgttcagctg cagcgtgatg
     6541 cacgaggccc tgcacaacca ctacacccag aagtccctga gcctgagccc cggcggcagc
     6601 ggtcttaatg atattttcga ggctcagaag attgagtggc atgaacacca ccaccatcac
     6661 cactaaggcc ggcctgtagt cactggcaca acgcgttacc cggtaagcca atcgggtata
     6721 cacggtcgtc atactgcaga cagggttctt ctactttgca agatagtctt gagtagtaaa
     6781 ataaatagat agagaaaatc agcctcgact gtgccttcta gttgccagcc atctgttgtt
     6841 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     6901 taaaatgagg aaattgcatc acaacactca accctatctc ggtctattct tttgatttat
     6961 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
     7021 acgcgaatta attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc
     7081 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc
     7141 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat
     7201 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
     7261 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga
     7321 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc
     7381 gggagcttgt atatccattt tcggatctga tcagcacgtg ttgacaatta atcatcggca
     7441 tagtatatcg gcatagtata atacgacaag gtgaggaact aaaccatgac cgagtacaag
     7501 cccacggtgc gcctcgccac ccgcgacgac gtccccaggg ccgtacgcac cctcgccgcc
     7561 gcgttcgccg actaccccgc cacgcgccac accgtcgatc cggaccgcca catcgagcgg
     7621 gtcaccgagc tgcaagaact cttcctcacg cgcgtcgggc tcgacatcgg caaggtgtgg
     7681 gtcgcggacg acggcgccgc ggtggcggtc tggaccacgc cggagagcgt cgaagcgggg
     7741 gcggtgttcg ccgagatcgg cccgcgcatg gccgagttga gcggttcccg gctggccgcg
     7801 cagcaacaga tggaaggcct cctggcgccg caccggccca aggagcccgc gtggttcctg
     7861 gccaccgtcg gcgtctcgcc cgaccaccag ggcaagggtc tgggcagcgc cgtcgtgctc
     7921 cccggagtgg aggcggccga gcgcgccggg gtgcccgcct tcctggagac atccgcgccc
     7981 cgcaacctcc ccttctacga gcggctcggc ttcaccgtca ccgccgacgt cgaggtgccc
     8041 gaaggaccgc gcacctggtg catgacccgc aagcccggtg cctgacacgt gctacgagat
     8101 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
     8161 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg
     8221 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     8281 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     8341 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcatt accaatgctt
     8401 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     8461 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gcgctgcgat
     8521 gataccgcga gaaccacgct caccggctcc ggatttatca gcaataaacc agccagccgg
     8581 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     8641 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     8701 cgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     8761 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     8821 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     8881 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     8941 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     9001 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     9061 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     9121 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     9181 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     9241 aatactcata ttcttcctt
//
