LOCUS       sequence-303145-        9046 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 9046)
  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 9046)
  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-CNTN3-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..9046
                     /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             6388..6432
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6433..6450
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6590..6607)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6596..6707
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6825..6845)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6830..7159
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7010..7145
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7072..7091
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7207..7254
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7273..7872
                     /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(7273..7292)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7769..7789
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    8002..8123
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8039..8058)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8093..8112
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(8176..8967,8968..9036))
                     /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     8799..8818
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(9037..9046,1..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 attgctgctg
     2761 caaggccctg tgttcatcaa agagcccagc aacagcatct tccccgtggg cagcgaggac
     2821 aagaagatca cactgcactg cgaggccaga ggcaacccat ctcctcacta tagatggcag
     2881 ctgaacggca gcgacatcga catgagcatg gaacaccggt acaagctgaa tggcggcaac
     2941 ctggtggtca tcaaccccaa cagaaactgg gacaccggca cctaccagtg cttcgccaca
     3001 aatagcctgg gcaccattgt gtccagagag gccaagctgc agttcgccta cctggaaaac
     3061 ttcaagacca agatgcggag caccgtgtcc gttagagaag gacaaggcgt ggtgctgctg
     3121 tgtggacctc ctcctcattc cggcgaactg tcctacgcct ggatcttcaa cgagtacccc
     3181 agcttcgtgg aagaggacag ccgcagattc gtgtcccaag agacaggcca cctgtacatc
     3241 agcaaggtgg aacccagcga cgtgggcaac tatacctgtg tggtcaccag catggtcacc
     3301 aacgccagag tgctgggatc tcctacacct ctggtgctga gatctgatgg cgtgatgggc
     3361 gagtacgagc ccaagatcga ggtgcagttt cctgaaacac tgcctgccgc caagggcagc
     3421 acagtgaagc tggaatgttt cgccctgggc aaccccattc cacagatcaa ttggcggaga
     3481 agcgacggcc tgcctttcag ctccaagatc aagctgagaa agttcagcgg cgtgctggaa
     3541 atccccaact tccagcaaga ggatgccggc agctatgagt gtatcgccga gaacagcaga
     3601 ggcaagaatg tggccagggg cagactgacc tactacgcca aacctcactg ggtgcagctg
     3661 atcaaggacg tggaaatcgc cgtggaagat agcctgtact gggagtgtag agccagcggc
     3721 aagcccaagc ctagctacag atggctgaaa aacggcgctg ccctggtgct ggaagagaga
     3781 acacagattg agaacggcgc cctgaccatc agcaacctgt ctgtgaccga tagcggcatg
     3841 ttccagtgca ttgccgaaaa caagcacggc ctggtgtaca gctctgccga actgaaagtg
     3901 gtggccagcg ctcccgactt cagcaagaac cccatgaaga aactggtgca ggtccaagtg
     3961 ggcagcctgg tgtctctgga ttgcaagcct agagcctctc ctagggctct gagcagctgg
     4021 aaaaaaggcg acgtgtccgt gcaagagcac gagagaatca gcctgctgaa cgacggcggc
     4081 ctgaagatcg ccaatgtgac aaaagccgac gccggcacat acacatgcat ggccgagaat
     4141 cagtttggca aggccaacgg cacaacccac ctggtcgtga ccgagcctac aagaatcaca
     4201 ctggccccta gcaacatgga tgtgtctgtg ggcgagagcg tgatcctgcc ttgtcaggtt
     4261 cagcacgacc ctctgctgga catcatcttc acctggtact tcaatggcgc cctggccgac
     4321 ttcaagaagg atggcagcca ctttgagaaa gtcggcggct ctagcagcgg cgacctgatg
     4381 atcagaaata tccagctgaa gcacagcggc aaatacgtgt gcatggtgca gacaggcgtg
     4441 gactctgtgt ctagcgccgc tgatctgatc gtcagaggat ctcccggacc tcctgagaac
     4501 gtgaaggtgg acgagatcac cgacacaaca gcccagctga gctggaaaga gggcaaagac
     4561 aatcacagcc ccgtgatcag ctacagcatc caggccagaa cacctttcag cgtcggctgg
     4621 cagacagtga ccacagtgcc tgaagtgatc gacggcaaga cccacacagc caccgtggtg
     4681 gaactgaacc cttgggtcga gtatgagttc cgcgtggtgg cctctaacaa gatcggaggc
     4741 ggagagccta gcctgcctag cgaaaaagtt cggactgaag aggccgtgcc tgaggtgcca
     4801 ccatctgaag ttaatggtgg cggcggaagc agaagcgagc tggttattac atgggacccc
     4861 gtgccagagg aactgcagaa tggcgaaggc tttggctacg tggtggcttt cagacctctg
     4921 ggcgtgacca cctggattca gacagtggtc acaagccccg acacacccag atacgtgttc
     4981 agaaacgaga gcatcgtgcc ttacagcccc tatgaagtga aagtgggcgt gtacaacaac
     5041 aaaggcgagg gcccttttag ccctgtgacc accgtgttta gcgccgaaga ggaacctaca
     5101 gtggcccctt ctcaggtgtc cgccaatagc ctgagcagca gcgagattga ggtgtcctgg
     5161 aacacaatcc cctggaagct gagcaacggc catctgctgg gatacgaagt gcggtattgg
     5221 aacggcggag gcaaagagga atccagcagc aagatgaagg tggccggcaa cgagacatct
     5281 gccagactga gaggcctgaa gtccaacctg gcctactata cagccgtgcg ggcctataat
     5341 tctgccggcg ctggaccatt cagcgccacc gtgaatgtga ccacaaagaa aacccctcca
     5401 agccagcctc ctggcaacgt cgtgtggaat gccaccgata ccaaagtgct gctgaactgg
     5461 gagcaagtga aggccatgga aaacgagtcc gaagtgaccg gctacaaagt gttctaccgg
     5521 accagcagcc agaacaacgt ccaggtgctg aacaccaaca agacctctgc agaactggtg
     5581 ctgcctatca aagaggacta catcatcgaa gtcaaggcca caacagatgg cggcgacggc
     5641 acatctagcg agcagatcag aatccccaga atcacctcca tggacgctgg cgcgccgggc
     5701 ggcgacaaga cccacacctg ccccccctgc ccagctccag aactgctggg agggccttcc
     5761 gtgttcctgt tcccccccaa gcccaaggac accctgatga tcagcaggac ccccgaggtg
     5821 acctgcgtgg tggtggccgt gtcccacgag gacccagagg tgaagttcaa ctggtacgtg
     5881 gacggcgtgg aggtgcacaa cgccaagacc aagcccagag aggagcagta caacagcacc
     5941 tacagggtgg tgtccgtgct gaccgtgctg caccaggact ggctgaacgg caaagaatac
     6001 aagtgcaaag tctccaacaa ggccctggct gccccaatcg aaaagacaat cagcaaggcc
     6061 aagggccagc cacgggagcc ccaggtgtac accctgcccc ccagccggga ggagatgacc
     6121 aagaaccagg tgtccctgac ctgtctggtg aagggcttct accccagcga tatcgccgtg
     6181 gagtgggaga gcaacggcca gcccgagaac aactacaaga ccaccccccc agtgctggac
     6241 agcgacggca gcttcttcct gtacagcaag ctgaccgtgg acaagtccag gtggcagcag
     6301 ggcaacgtgt tcagctgcag cgtgatgcac gaggccctgc acaaccacta cacccagaag
     6361 tccctgagcc tgagccccgg cggcagcggt cttaatgata ttttcgaggc tcagaagatt
     6421 gagtggcatg aacaccacca ccatcaccac taaggccggc ctgtagtcac tggcacaacg
     6481 cgttacccgg taagccaatc gggtatacac ggtcgtcata ctgcagacag ggttcttcta
     6541 ctttgcaaga tagtcttgag tagtaaaata aatagataga gaaaatcagc ctcgactgtg
     6601 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
     6661 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcaca acactcaacc
     6721 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     6781 aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt
     6841 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     6901 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     6961 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     7021 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     7081 agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg
     7141 aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca
     7201 gcacgtgttg acaattaatc atcggcatag tatatcggca tagtataata cgacaaggtg
     7261 aggaactaaa ccatgaccga gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc
     7321 cccagggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac gcgccacacc
     7381 gtcgatccgg accgccacat cgagcgggtc accgagctgc aagaactctt cctcacgcgc
     7441 gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg
     7501 accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc gcgcatggcc
     7561 gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct ggcgccgcac
     7621 cggcccaagg agcccgcgtg gttcctggcc accgtcggcg tctcgcccga ccaccagggc
     7681 aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg cgccggggtg
     7741 cccgccttcc tggagacatc cgcgccccgc aacctcccct tctacgagcg gctcggcttc
     7801 accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag
     7861 cccggtgcct gacacgtgct acgagatttc gattccaccg ccgccttcta tgaaaggttg
     7921 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     7981 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
     8041 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     8101 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
     8161 gcgtaatcat ggtcattacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     8221 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     8281 cttaccatct ggccccagcg ctgcgatgat accgcgagaa ccacgctcac cggctccgga
     8341 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     8401 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     8461 taatagtttg cgcaacgttg ttgccatcgc tacaggcatc gtggtgtcac gctcgtcgtt
     8521 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     8581 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     8641 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     8701 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     8761 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     8821 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     8881 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     8941 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     9001 gggaataagg gcgacacgga aatgttgaat actcatattc ttcctt
//
