LOCUS       sequence-303123-        8920 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 8920)
  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 8920)
  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-CDON-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..8920
                     /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             6262..6306
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6307..6324
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6464..6481)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6470..6581
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6699..6719)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6704..7033
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6884..7019
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6946..6965
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7081..7128
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7147..7746
                     /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(7147..7166)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7643..7663
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7876..7997
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7913..7932)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7967..7986
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(8050..8841,8842..8910))
                     /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     8673..8692
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8911..8920,1..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 tctggcccct
     2761 tactttacat ctgaacctct gagcgccgtg cagaaactcg gaggacctgt ggttctgcac
     2821 tgtagcgctc agcctgtgac caccagaatc agctggctgc acaacggcaa gaccctggac
     2881 ggaaacctgg aacacgtgaa gatccaccag ggcaccctga ccatcctgag cctgaatagc
     2941 agcctgctgg gctactacca gtgcctggcc aacaattcta tcggcgccat tgtgtctggc
     3001 cctgccacag tgtctgtggc cgttctgggc gattttggca gcagcaccaa acacgtgatc
     3061 accgccgagg aaaagagcgc cggctttatc ggatgtagag tgcccgagag caaccccaag
     3121 gccgaagtgc ggtataagat cagaggcaag tggctggaac acagcaccga gaactacctg
     3181 atcctgccaa gcggcaacct gcagatcctg aacgtgtccc tggaagataa gggcagctac
     3241 aagtgcgccg cctacaatcc agtgacacac cagctgaagg tggaacccat cggcagaaag
     3301 ctgctggtgt ccagacctag cagcgacgac gtgcacattc tgcaccctac acattctcag
     3361 gccctggccg tgctgagcag atctcctgtt acactggaat gcgtggtgtc cggcgttcca
     3421 gctcctcagg tgtactggct gaaggacgga caggatatcg cccctggcag caattggaga
     3481 aggctgtact ctcacctggc caccgattct gtggaccctg ccgatagcgg caattactcc
     3541 tgtatggccg gcaacaagag cggcgacgtg aaatacgtga cctacatggt caacgtgctg
     3601 gaacatgcca gcatcagcaa gggcctgcag gaccagattg tgtctctggg cgccaccgtg
     3661 cacttcacct gtgatgtgca tggcaacccc gctcctaact gcacctggtt tcacaacgcc
     3721 cagcctattc accctagcgc cagacatctg acagccggca atggactgaa gatcagcggc
     3781 gtgacagtgg aagatgtggg catgtaccag tgtgtggccg acaacggcat cggctttatg
     3841 cactctaccg gcagactgga aatcgagaac gacggcggct tcaagcccgt gatcattaca
     3901 gcccctgtgt ccgccaaagt ggccgatggg gattttgtga ccctgagctg taatgcctct
     3961 ggcctgcctg tgcctgtgat ccgttggtac gatagccacg gcctgatcac cagccatcct
     4021 agccaggtgc tgcggagcaa gagcagaaag tctcagctgt caagacccga gggcctgaat
     4081 ctggaacccg tgtactttgt tctgagccag gctggcgcct ctagcctgca cattcaggct
     4141 gtgacacaag agcacgccgg caagtatatc tgcgaggccg ccaatgagca cggcacaact
     4201 caagccgaag ccagcctgat ggtggtgccc ttcgagacaa acacaaaggc cgagacagtg
     4261 accctgcctg acgccgctca gaacgacgac agatctaaga gagatggcag cgagacaggc
     4321 ctgctgtcca gctttcctgt gaaagtgcac cccagcgccg tggaatctgc ccctgagaaa
     4381 aatgccagcg gcatcagcgt gcccgacgct cctattattc tgagccctcc tcagacacac
     4441 acccctgaca cctacaacct cgtttggaga gccggaaagg atggcggcct gcctatcaac
     4501 gcctacttcg tgaagtaccg gaagctggac gatggcgtgg gaatgctcgg atcttggcac
     4561 acagtcagag tgccaggctc cgagaatgag ctgcatctgg ccgaactgga accttccagc
     4621 ctgtacgagg tgctgatggt tgctagatct gctgccggcg aaggacagcc tgccatgctg
     4681 accttccgga ccagcaaaga gaaaaccgcc agcagcaaga acacccaggc ctcttctcca
     4741 cctgtgggca tccctaagta ccccgtggtg tctgaagccg ctaacaacaa ctttggcgtg
     4801 gtgctgaccg acagcagcag acatagcgga gtgcctgagg ctcccgatag acctacaatc
     4861 agcaccgcca gcgaaaccag cgtgtacgtg acatggattc ccagagccaa tggcggcagc
     4921 cctatcaccg ccttcaaggt cgagtacaag cggatgagaa ccagcaactg gctggtggcc
     4981 gccgaagata ttcctcctag caagctgagc gtggaagtgc gctctctgga accaggcagc
     5041 acctacaagt tcagagtgat cgccatcaac cactacggcg agagcttcag atccagcgcc
     5101 agcagacctt accaggtcgt gggcttcccc aacagattca gcagcaggcc aatcacaggc
     5161 cctcacattg cctacaccga ggccgtgtct gacacccaga tcatgctgaa gtggacctac
     5221 attcccagct ccaacaacaa cacccctatc cagggcttct acatctacta cagacccacc
     5281 gacagcgaca acgactccga ctacaagcga gatgtggtgg aaggcagcaa gcagtggcac
     5341 atgattggcc atctgcagcc cgagacaagc tacgacatca agatgcagtg cttcaacgaa
     5401 ggcggcgagt ccgagttcag caacgtgatg atctgcgaga caaaagtgaa gagagtgcct
     5461 ggggccagcg agtaccctgt gaaggatctg agcacccctc ctaattctct cggctctggc
     5521 ggaaatgtgg gcccagctac atctcctgcc agatcctctg atggcgcgcc gggcggcgac
     5581 aagacccaca cctgcccccc ctgcccagct ccagaactgc tgggagggcc ttccgtgttc
     5641 ctgttccccc ccaagcccaa ggacaccctg atgatcagca ggacccccga ggtgacctgc
     5701 gtggtggtgg ccgtgtccca cgaggaccca gaggtgaagt tcaactggta cgtggacggc
     5761 gtggaggtgc acaacgccaa gaccaagccc agagaggagc agtacaacag cacctacagg
     5821 gtggtgtccg tgctgaccgt gctgcaccag gactggctga acggcaaaga atacaagtgc
     5881 aaagtctcca acaaggccct ggctgcccca atcgaaaaga caatcagcaa ggccaagggc
     5941 cagccacggg agccccaggt gtacaccctg ccccccagcc gggaggagat gaccaagaac
     6001 caggtgtccc tgacctgtct ggtgaagggc ttctacccca gcgatatcgc cgtggagtgg
     6061 gagagcaacg gccagcccga gaacaactac aagaccaccc ccccagtgct ggacagcgac
     6121 ggcagcttct tcctgtacag caagctgacc gtggacaagt ccaggtggca gcagggcaac
     6181 gtgttcagct gcagcgtgat gcacgaggcc ctgcacaacc actacaccca gaagtccctg
     6241 agcctgagcc ccggcggcag cggtcttaat gatattttcg aggctcagaa gattgagtgg
     6301 catgaacacc accaccatca ccactaaggc cggcctgtag tcactggcac aacgcgttac
     6361 ccggtaagcc aatcgggtat acacggtcgt catactgcag acagggttct tctactttgc
     6421 aagatagtct tgagtagtaa aataaataga tagagaaaat cagcctcgac tgtgccttct
     6481 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     6541 actcccactg tcctttccta ataaaatgag gaaattgcat cacaacactc aaccctatct
     6601 cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg
     6661 agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg
     6721 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     6781 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     6841 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     6901 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     6961 cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     7021 aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg atcagcacgt
     7081 gttgacaatt aatcatcggc atagtatatc ggcatagtat aatacgacaa ggtgaggaac
     7141 taaaccatga ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga cgtccccagg
     7201 gccgtacgca ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat
     7261 ccggaccgcc acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac gcgcgtcggg
     7321 ctcgacatcg gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt ctggaccacg
     7381 ccggagagcg tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat ggccgagttg
     7441 agcggttccc ggctggccgc gcagcaacag atggaaggcc tcctggcgcc gcaccggccc
     7501 aaggagcccg cgtggttcct ggccaccgtc ggcgtctcgc ccgaccacca gggcaagggt
     7561 ctgggcagcg ccgtcgtgct ccccggagtg gaggcggccg agcgcgccgg ggtgcccgcc
     7621 ttcctggaga catccgcgcc ccgcaacctc cccttctacg agcggctcgg cttcaccgtc
     7681 accgccgacg tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg caagcccggt
     7741 gcctgacacg tgctacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     7801 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     7861 ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     7921 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     7981 ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa
     8041 tcatggtcat taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     8101 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     8161 atctggcccc agcgctgcga tgataccgcg agaaccacgc tcaccggctc cggatttatc
     8221 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     8281 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     8341 tttgcgcaac gttgttgcca tcgctacagg catcgtggtg tcacgctcgt cgtttggtat
     8401 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     8461 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     8521 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     8581 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     8641 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     8701 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     8761 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     8821 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     8881 aagggcgaca cggaaatgtt gaatactcat attcttcctt
//
