LOCUS       sequence-303143-        9043 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 9043)
  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 9043)
  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-CNTN4-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..9043
                     /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             6385..6429
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6430..6447
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6587..6604)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6593..6704
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6822..6842)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6827..7156
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7007..7142
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7069..7088
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7204..7251
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7270..7869
                     /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(7270..7289)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7766..7786
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7999..8120
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8036..8055)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8090..8109
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(8173..8964,8965..9033))
                     /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     8796..8815
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(9034..9043,1..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 tgattctaca
     2761 ctgcacggcc ccatcttcat ccaagagcct tctccagtga tgttccctct ggacagcgaa
     2821 gagaagaaag tgaagctgaa ctgcgaagtg aagggcaacc ccaagcctca catccggtgg
     2881 aagctgaatg gcaccgatgt ggacaccggc atggacttca gatacagcgt ggtggaaggc
     2941 agcctgctga tcaacaaccc caacaagaca caggacgccg gcacctacca gtgtaccgcc
     3001 acaaatagct tcggcaccat cgtgtccaga gaggccaagc tgcagttcgc ctacctggac
     3061 aacttcaaga cccggaccag atccaccgtg tccgttagaa gaggacaagg catggtgctg
     3121 ctgtgtggcc ctcctcctca ttccggcgaa ctgtcctacg cctggatctt caacgagtac
     3181 cccagctacc aggacaaccg cagattcgtg tcccaagaga caggcaacct gtatatcgcc
     3241 aaggtggaaa agtccgacgt gggcaactac acctgtgtgg tcaccaacac cgtgaccaac
     3301 cacaaggtgc tgggacctcc aacacctctg atcctgagaa acgatggcgt gatgggcgag
     3361 tacgagccca agatcgaggt gcagtttccc gagacagtgc ctacagccaa gggcgccaca
     3421 gtgaaactgg aatgctttgc cctgggcaac cctgtgccta ccattatctg gcggagagcc
     3481 gatggcaagc ctatcgccag aaaggccaga aggcacaaga gcaacggcat cctggaaatc
     3541 cccaacttcc agcaagagga tgccggcctg tatgagtgcg tggccgagaa cagcagaggc
     3601 aagaatgtgg ccagaggcca gctgaccttc tacgcccagc ctaactggat tcagaagatc
     3661 aacgacatcc acgtggccat ggaagagaac gtgttctggg agtgcaaggc caacggcaga
     3721 cccaagccaa cctacaagtg gctgaagaat ggcgagcccc tgctgaccag agacagaatc
     3781 cagatcgagc agggcaccct gaacatcaca atcgtgaatc tgagcgacgc cgggatgtac
     3841 cagtgtctgg ccgaaaacaa gcacggcgtg atcttcagca atgccgagct gtctgtgatc
     3901 gccgtgggac ccgatttcag cagaacactg ctgaagagag tgaccctggt caaagtcggc
     3961 ggcgaggttg tgatcgagtg caagcctaag gcctctccta agcctgtgta cacctggaag
     4021 aagggcagag acatcctgaa agagaacgag cggatcacca tcagcgagga cggcaacctg
     4081 agaatcatca acgtgaccaa gagcgacgct ggcagctata cctgtatcgc caccaatcac
     4141 tttggcaccg ccagcagcac aggcaatctg gtggtcaagg accccaccag agtgatggtg
     4201 cctccaagca gcatggatgt gacagtgggc gagagcatcg tgctgccttg tcaagtgaca
     4261 cacgaccaca gcctggacat cgtgttcacc tggtccttca acggccacct gatcgacttc
     4321 gacagagatg gcgaccactt cgagagagtc ggaggacagg attctgccgg cgacctgatg
     4381 atccggaaca tccagctgaa gcacgccggc aaatacgtgt gcatggtgca gacctccgtg
     4441 gacagactgt ctgccgctgc cgatctgatc gttagaggac ctcctggacc tccagaggcc
     4501 gtgaccatcg atgagatcac cgatacaaca gcccagctgt cttggcggcc tggacctgat
     4561 aatcacagcc ccatcactat gtacgtgatc caggccagaa cacccttcag cgttggatgg
     4621 caggctgtgt ctacagtgcc tgagctgatc gacggcaaga cctttacagc caccgtcgtg
     4681 ggcctgaatc cttgggtcga gtatgagttt agaaccgtgg ccgccaacgt gatcggaatc
     4741 ggagaaccta gcagacccag cgagaagcgg agaactgaag aggccctgcc tgaagtgacc
     4801 cctgctaatg tttctggcgg cggaggctct aagtccgagc tggttattac ctgggagact
     4861 gtgcccgagg aactgcagaa tggcagaggc tttggctacg tggtggcctt cagaccctac
     4921 ggcaagatga tctggatgct gaccgtgctg gcttccgccg atgccagcag atacgtgttc
     4981 agaaacgaga gcgtgcaccc tttcagcccc ttcgaagtga aagtgggcgt gttcaacaac
     5041 aaaggcgagg gcccttttag ccccaccaca gtggtttaca gcgccgagga agaacctacc
     5101 aagcctcctg ccagcatctt cgccagaagc ctgtctgcca ccgacatcga agtgttttgg
     5161 gcctctccac tggaaaagaa ccggggaaga atccagggct atgaagtgaa gtactggcgg
     5221 cacgaggaca aagaggaaaa cgcccggaag atccggaccg tgggcaatca gaccagcacc
     5281 aagatcacca atctgaaggg cagcgtgctg taccacctgg ccgtgaaggc ctataatagc
     5341 gccggaacag gaccaagcag cgccaccgtg aatgtgacca caagaaagcc tccacctagc
     5401 cagcctccag gcaacatcat ctggaacagc agcgacagca agatcatcct gaactgggac
     5461 caagtgaagg ccctggacaa tgagtccgaa gtcaagggct ataaggtgct gtacagatgg
     5521 aaccggcagt ccagcaccag cgtgatcgag acaaacaaga ccagcgtgga actgagcctg
     5581 cctttcgacg aggactacat catcgagatc aagccctttt ccgatggcgg cgacggaagc
     5641 agctctgagc agatcagaat ccccaagatc agcaacgcct acgctggcgc gccgggcggc
     5701 gacaagaccc acacctgccc cccctgccca gctccagaac tgctgggagg gccttccgtg
     5761 ttcctgttcc cccccaagcc caaggacacc ctgatgatca gcaggacccc cgaggtgacc
     5821 tgcgtggtgg tggccgtgtc ccacgaggac ccagaggtga agttcaactg gtacgtggac
     5881 ggcgtggagg tgcacaacgc caagaccaag cccagagagg agcagtacaa cagcacctac
     5941 agggtggtgt ccgtgctgac cgtgctgcac caggactggc tgaacggcaa agaatacaag
     6001 tgcaaagtct ccaacaaggc cctggctgcc ccaatcgaaa agacaatcag caaggccaag
     6061 ggccagccac gggagcccca ggtgtacacc ctgcccccca gccgggagga gatgaccaag
     6121 aaccaggtgt ccctgacctg tctggtgaag ggcttctacc ccagcgatat cgccgtggag
     6181 tgggagagca acggccagcc cgagaacaac tacaagacca cccccccagt gctggacagc
     6241 gacggcagct tcttcctgta cagcaagctg accgtggaca agtccaggtg gcagcagggc
     6301 aacgtgttca gctgcagcgt gatgcacgag gccctgcaca accactacac ccagaagtcc
     6361 ctgagcctga gccccggcgg cagcggtctt aatgatattt tcgaggctca gaagattgag
     6421 tggcatgaac accaccacca tcaccactaa ggccggcctg tagtcactgg cacaacgcgt
     6481 tacccggtaa gccaatcggg tatacacggt cgtcatactg cagacagggt tcttctactt
     6541 tgcaagatag tcttgagtag taaaataaat agatagagaa aatcagcctc gactgtgcct
     6601 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt
     6661 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcacaaca ctcaacccta
     6721 tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa
     6781 atgagctgat ttaacaaaaa tttaacgcga attaattctg tggaatgtgt gtcagttagg
     6841 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     6901 gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     6961 gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac
     7021 tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga
     7081 ggccgaggcc gcctctgcct ctgagctatt ccagaagtag tgaggaggct tttttggagg
     7141 cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc attttcggat ctgatcagca
     7201 cgtgttgaca attaatcatc ggcatagtat atcggcatag tataatacga caaggtgagg
     7261 aactaaacca tgaccgagta caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc
     7321 agggccgtac gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc
     7381 gatccggacc gccacatcga gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc
     7441 gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc
     7501 acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg catggccgag
     7561 ttgagcggtt cccggctggc cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg
     7621 cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct cgcccgacca ccagggcaag
     7681 ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc
     7741 gccttcctgg agacatccgc gccccgcaac ctccccttct acgagcggct cggcttcacc
     7801 gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc
     7861 ggtgcctgac acgtgctacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     7921 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     7981 gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     8041 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     8101 aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg
     8161 taatcatggt cattaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     8221 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     8281 accatctggc cccagcgctg cgatgatacc gcgagaacca cgctcaccgg ctccggattt
     8341 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     8401 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     8461 tagtttgcgc aacgttgttg ccatcgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     8521 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     8581 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     8641 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     8701 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     8761 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     8821 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     8881 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     8941 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     9001 aataagggcg acacggaaat gttgaatact catattcttc ctt
//
