LOCUS       sequence-303165-        9280 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 9280)
  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 9280)
  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-CHL1-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..9280
                     /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             6622..6666
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6667..6684
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6824..6841)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6830..6941
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(7059..7079)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        7064..7393
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7244..7379
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7306..7325
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7441..7488
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7507..8106
                     /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(7507..7526)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     8003..8023
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    8236..8357
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8273..8292)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8327..8346
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(8410..9201,9202..9270))
                     /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     9033..9052
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(9271..9280,1..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 cggccgccat tgagattcct
     2761 agctctgtgc agcaggtccc caccatcatc aagcagagca aggtgcaggt cgcctttcca
     2821 ttcgacgagt acttccagat cgagtgcgag gccaagggca accctgagcc tacctttagc
     2881 tggaccaagg acggcaaccc cttctacttc accgaccaca gaatcatccc cagcaacaac
     2941 agcggcacct tcagaatccc caacgagggc cacatcagcc acttccaggg caagtacaga
     3001 tgcttcgcct ccaacaagct gggaatcgcc atgagcgaag agatcgagtt catcgtgccc
     3061 agcgtgccca agtttcccaa agagaagatc gaccctctcg aggtggaaga gggcgaccct
     3121 attgtgctgc cctgcaatcc tcctaagggc ctgcctcctc tgcacatcta ctggatgaac
     3181 atcgagctgg aacacatcga gcaggacgag cgggtgtaca tgagccagaa gggcgacctg
     3241 tacttcgcca acgtggaaga gaaggacagc cggaacgact actgctgctt cgccgccttt
     3301 cctagactgc ggaccatcgt gcagaaaatg cccatgaagc tgaccgtgaa cagcagcaac
     3361 tctatcaagc agcggaagcc caagctgctg ctgcctccta cagagagcgg aagcgagagc
     3421 agcatcacca tcctgaaggg cgagatcctg ctgctggaat gctttgccga gggactgcct
     3481 acacctcagg tggactggaa caagatcggc ggcgatctgc ctaagggcag agagacaaaa
     3541 gagaactacg gcaagaccct gaagatcgag aacgtgtcct accaggacaa gggcaattac
     3601 cggtgtaccg ccagcaactt cctgggcaca gccacacacg acttccacgt gatcgtggaa
     3661 gaacctcctc ggtggaccaa gaaacctcag agcgccgtgt acagcaccgg cagcaatgga
     3721 attctgctgt gcgaagccga gggcgagcct cagccaacaa tcaagtggcg cgtgaacggc
     3781 agccccgtgg acaatcatcc ttttgctggc gacgtggtgt tccccagaga gatcagcttc
     3841 accaacctgc agcctaacca cacagccgtg taccagtgtg aagccagcaa tgtgcacggc
     3901 acaatcctgg ccaacgccaa cattgacgtg gtggatgtgc ggcccctgat ccagacaaag
     3961 gacggcgaga attacgccac cgtcgtgggc tactctgcct tcctgcactg cgagttcttc
     4021 gcctctcctg aggctgtggt gtcctggcag aaagtggaag aagtgaagcc cctcgaaggc
     4081 cgcagatacc acatctacga gaacggcacc ctgcagatca accggaccac agaggaagat
     4141 gccggcagct actcttgctg ggtcgagaat gccatcggca agacagccgt gaccgccaac
     4201 ctggacatca gaaacgccac caagctgcgg gtgtccccaa agaaccccag aattcctaag
     4261 ctgcacatgc tggaactgca ttgcgagagc aagtgcgaca gccacctgaa gcacagcctg
     4321 aagctgtcct ggtccaagga tggcgaggcc tttgagatca acggcaccga ggatggccgg
     4381 atcatcatcg atggcgccaa tctgaccatc agcaacgtga ccctcgagga ccagggcatc
     4441 tactgttgta gcgcccacac tgccctggac tctgccgccg atatcaccca agtgaccgtg
     4501 ctggatgtcc ccgatcctcc tgagaatctg cacctgagcg agcggcagaa tagaagcgtc
     4561 agactgacat gggaggctgg cgccgaccac aacagcaaca tctccgagta catcgtggaa
     4621 ttcgagggca acaaagagga acccggccgt tgggaagaac tgaccagagt gcagggaaag
     4681 aaaaccaccg tgatcctgcc tctggctccc ttcgtgcgct accagtttag agtgatcgcc
     4741 gtgaacgaag tgggcagaag ccagccttct cagcccagcg atcaccacga aacacctcca
     4801 gccgctccag acagaaaccc tcagaacatc cgcgtgcagg ccagccagcc taaagaaatg
     4861 atcatcaagt gggagcccct gaagtccatg gaacagaatg gccctgggct cgagtacaga
     4921 gtgacctgga aacctcaagg cgcccctgtg gaatgggaag aagagacagt caccaatcac
     4981 accctgagag tgatgacccc tgccgtgtac gccccttacg atgtgaaagt gcaggctatc
     5041 aaccagctcg gcagcggccc agatcctcag tctgtgacac tgtacagcgg cgaggactac
     5101 cccgatacag cccctgtgat tcatggcgtg gacgtgatca acagcaccct ggtcaaagtg
     5161 acatggtcca cagtgcccaa ggacagagtg cacggtagac tgaagggcta ccagatcaac
     5221 tggtggaaaa caaagagcct gctggacggc cggacacacc ctaaagaagt caacatcctg
     5281 cggttcagcg gccagagaaa ctccggcatg gtgcctagcc tggatgcctt cagcgagttc
     5341 cacctgacag tgctggccta caactccaaa ggcgccggac ctgagagcga gccctacatc
     5401 tttcagacac ctgagggcgt gccagagcag cccacctttc tgaaagtgat caaggtggac
     5461 aaggacaccg ccacactgtc ttggggcctg ccaaagaagc tgaacggcaa cctgacaggc
     5521 tatctgctgc agtatcagat catcaacgac acctacgaga tcggcgagct gaacgacatc
     5581 aacatcacca cacctagcaa gcccagctgg cacctgtcca acctgaatgc caccaccaag
     5641 tacaagttct acctgcgggc ctgcaccagc caaggctgtg gaaagcctat caccgaggaa
     5701 agcagcacac tcggcgaggg ctctaaaggc atcggaaaga tcagcggcgt gaacctgaca
     5761 cagaaaacac accccatcga ggtgttcgag ccaggcgccg aacacattgt gcggctgatg
     5821 accaagaact ggggcgacaa cgacagcatc ttccaggatg tgatcgagac aagaggcaga
     5881 gagtacgccg gcctgtacga cgacatcagc acacaaggat ggggcgcgcc gggcggcgac
     5941 aagacccaca cctgcccccc ctgcccagct ccagaactgc tgggagggcc ttccgtgttc
     6001 ctgttccccc ccaagcccaa ggacaccctg atgatcagca ggacccccga ggtgacctgc
     6061 gtggtggtgg ccgtgtccca cgaggaccca gaggtgaagt tcaactggta cgtggacggc
     6121 gtggaggtgc acaacgccaa gaccaagccc agagaggagc agtacaacag cacctacagg
     6181 gtggtgtccg tgctgaccgt gctgcaccag gactggctga acggcaaaga atacaagtgc
     6241 aaagtctcca acaaggccct ggctgcccca atcgaaaaga caatcagcaa ggccaagggc
     6301 cagccacggg agccccaggt gtacaccctg ccccccagcc gggaggagat gaccaagaac
     6361 caggtgtccc tgacctgtct ggtgaagggc ttctacccca gcgatatcgc cgtggagtgg
     6421 gagagcaacg gccagcccga gaacaactac aagaccaccc ccccagtgct ggacagcgac
     6481 ggcagcttct tcctgtacag caagctgacc gtggacaagt ccaggtggca gcagggcaac
     6541 gtgttcagct gcagcgtgat gcacgaggcc ctgcacaacc actacaccca gaagtccctg
     6601 agcctgagcc ccggcggcag cggtcttaat gatattttcg aggctcagaa gattgagtgg
     6661 catgaacacc accaccatca ccactaaggc cggcctgtag tcactggcac aacgcgttac
     6721 ccggtaagcc aatcgggtat acacggtcgt catactgcag acagggttct tctactttgc
     6781 aagatagtct tgagtagtaa aataaataga tagagaaaat cagcctcgac tgtgccttct
     6841 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     6901 actcccactg tcctttccta ataaaatgag gaaattgcat cacaacactc aaccctatct
     6961 cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg
     7021 agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg
     7081 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     7141 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     7201 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     7261 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     7321 cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     7381 aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg atcagcacgt
     7441 gttgacaatt aatcatcggc atagtatatc ggcatagtat aatacgacaa ggtgaggaac
     7501 taaaccatga ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga cgtccccagg
     7561 gccgtacgca ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat
     7621 ccggaccgcc acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac gcgcgtcggg
     7681 ctcgacatcg gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt ctggaccacg
     7741 ccggagagcg tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat ggccgagttg
     7801 agcggttccc ggctggccgc gcagcaacag atggaaggcc tcctggcgcc gcaccggccc
     7861 aaggagcccg cgtggttcct ggccaccgtc ggcgtctcgc ccgaccacca gggcaagggt
     7921 ctgggcagcg ccgtcgtgct ccccggagtg gaggcggccg agcgcgccgg ggtgcccgcc
     7981 ttcctggaga catccgcgcc ccgcaacctc cccttctacg agcggctcgg cttcaccgtc
     8041 accgccgacg tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg caagcccggt
     8101 gcctgacacg tgctacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     8161 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     8221 ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     8281 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     8341 ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa
     8401 tcatggtcat taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     8461 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     8521 atctggcccc agcgctgcga tgataccgcg agaaccacgc tcaccggctc cggatttatc
     8581 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     8641 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     8701 tttgcgcaac gttgttgcca tcgctacagg catcgtggtg tcacgctcgt cgtttggtat
     8761 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     8821 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     8881 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     8941 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     9001 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     9061 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     9121 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     9181 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     9241 aagggcgaca cggaaatgtt gaatactcat attcttcctt
//
