LOCUS       sequence-444815-        8605 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 8605)
  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 8605)
  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-TAPBPL-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..8605
                     /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        3105..3484
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3485..3688
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3639..3659
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     3685..3709
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        3919..3937
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     3919..3936
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     3950..3969
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             5947..5991
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5992..6009
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6149..6166)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6155..6266
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6384..6404)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6389..6718
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6569..6704
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6631..6650
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6766..6813
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6832..7431
                     /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(6832..6851)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7328..7348
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7561..7682
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7598..7617)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7652..7671
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7735..8526,8527..8595))
                     /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     8358..8377
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8596..8605,1..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 gctgatgaat cccctaatga tttttatcaa aatcattaag ttaaggtaga tacacatctt
     1681 gtcatatgat caaatggttt cgccaaaaat caataatcag acaacaaaat gtgcgaactc
     1741 gatattttac acgactctct ttaccaattc tgccccgaat tacacttaaa acgactcaac
     1801 agcttaacgt tggcttgcca cgccttactt gactgtaaaa ctctcactct taccgaactt
     1861 ggccgtaacc tgccaaccaa agcgagaaca aaacataaca tcaaacgaat cgaccgattg
     1921 ttaggtaatc gtcacctcca caaagagcga ctcgctgtat accgttggca tgctagcttt
     1981 atctgttcgg gcaatacgat gcccattgta cttgttgact ggtctgatat ccgtgagcaa
     2041 aaacggctta tggtattgcg agcttcagtc gcactacacg gtcgttctgt tactctttat
     2101 gagaaagcgt tcccgctttc agagcaatgt tcaaagaaag ctcatgacca atttctagcc
     2161 gaccttgcga gcattctacc gagtaacacc acaccgctca ttgtcagtga tgctggcttt
     2221 aaagtgccat ggtataaatc cgttgagaag ctgggttggt actggttaag tcgagtaaga
     2281 ggaaaagtac aatatgcaga cctaggagcg gaaaactgga aacctatcag caacttacat
     2341 gatatgtcat ctagtcactc aaagacttta ggctataaga ggctgactaa aagcaatcca
     2401 atctcatgcc aaattctatt gtataaatct cgctctaaag gccgaaaaaa tcagcgctcg
     2461 acacggactc attgtcacca cccgtcacct aaaatctact cagcgtcggc aaaggagcca
     2521 tggattctag caactaactt acctgttgaa attcgaacac ccaaacaact tgttaatatc
     2581 tattcgaagc gaatgcagat tgaagaaacc ttccgagact tgaaaagtcc tgcctacgga
     2641 ctaggcctac gccatagccg aacgagcagc tcagagcgtt ttgatatcat gctgctaatc
     2701 gccctgatgc ttcaactaac atgttggctt gcgggcgttc atgctcagaa acaaggttgg
     2761 gacaagcact tccaggctaa cacagtcaga aatcgaaacg tactctcaac agttcgctta
     2821 ggcatggaag ttttgcggca ttctggctac acaataacaa gggaagactc actcgtggct
     2881 gcaaccctgc ttactcaaaa tctattcaca catggttacg ttttggggaa attatgaggg
     2941 gatctctcag tgcacagtgg cgttgactca atattggcca ttagccatat tattcattgg
     3001 ttatatagca taaatcaata ttggctattg gccattgcat acgttgtatc tatatcataa
     3061 tatgtacatt tatattggct catgtccaat atgaccgcca tgttggcatt gattattgac
     3121 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
     3181 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
     3241 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
     3301 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
     3361 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
     3421 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
     3481 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
     3541 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
     3601 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
     3661 acggtgggag gtctatataa gcagagctcg tttagtgaac cgtcagatcg cctggagacg
     3721 ccatccacgc tgttttgacc tccatagaag acaccgggac cgatccagcc tccgcggccg
     3781 ggaacggtgc attggaacgc ggattccccg tgccaagagt gacgtaagta ccgcctatag
     3841 agtctatagg cccaccccct tggcttcgtt agaacgcggc tacaattaat acataacctt
     3901 atgtatcata cacatacgat ttaggtgaca ctatagaata acatccactt tgcctttctc
     3961 tccacaggtg tccactccca ggtccaactg cacctcggtt ctatcgaaaa cgcgcctgct
     4021 agcgccacca tgaagaccat cattgccctg agctacatct tctgcctggt gttcgctgcg
     4081 gccgccgctg aaacaaaacc tcatcctgct gaaggccagt ggcgcgctgt ggatgttgtg
     4141 ctggattgct tcctggccaa ggacggcgct catagaggtg ctctggccag ctctgaggat
     4201 agagccagag cttccctggt gctgaaacag gtgcccgtgc tggatgatgg cagcctggaa
     4261 gatttcaccg acttccaagg cggcacactg gcccaggatg accctcctat catcttcgag
     4321 gccagcgtgg acctggtgca gattcctcaa gctgaagccc tgctgcacgc cgactgtagc
     4381 ggaaaagaag tgacctgcga gatcagccgg tacttcctgc agatgaccga gacaaccgtg
     4441 aaaaccgccg cctggttcat ggccaacatg caggtttcag gcggaggccc tagcatcagc
     4501 ctggtcatga agacccctag agtgaccaag aacgaggccc tgtggcaccc cacactgaac
     4561 ttgcctttga gccctcaagg caccgtgcgg accgccgtgg aatttcaagt gatgacacag
     4621 acccagagcc tgtccttcct gctgggatct agcgcctctc tggattgcgg cttttctatg
     4681 gcccctggcc tggacctgat cagcgttgaa tggcggctgc agcacaaagg cagaggccag
     4741 ctggtgtact cttggacagc tggacagggc caagccgtca gaaaaggcgc tacactggaa
     4801 cctgctcagc tcggcatggc cagagatgct tctctgacac tgcctggcct gaccatccag
     4861 gacgagggca cctacatctg ccagatcacc accagcctgt acagagccca gcagattatc
     4921 cagctgaaca tccaggcctc tccaaaagtg cggctgagcc tggccaatga ggctctgctg
     4981 cctacactga tctgcgacat tgccggctac taccctctgg acgtggtggt cacatggacc
     5041 agagaggaac tcggcggctc tcctgctcaa gtgtctggcg cctctttcag cagcctgaga
     5101 cagtctgtgg ccggcaccta ctctatcagc agctctctga cagccgagcc tggatctgct
     5161 ggcgccacct acacatgtca agtgacccac atctccctgg aagaacccct gggagcctct
     5221 acacaggtgg tgcctccaga gagaagaggc gcgccgggcg gcgacaagac ccacacctgc
     5281 cccccctgcc cagctccaga actgctggga gggccttccg tgttcctgtt cccccccaag
     5341 cccaaggaca ccctgatgat cagcaggacc cccgaggtga cctgcgtggt ggtggccgtg
     5401 tcccacgagg acccagaggt gaagttcaac tggtacgtgg acggcgtgga ggtgcacaac
     5461 gccaagacca agcccagaga ggagcagtac aacagcacct acagggtggt gtccgtgctg
     5521 accgtgctgc accaggactg gctgaacggc aaagaataca agtgcaaagt ctccaacaag
     5581 gccctggctg ccccaatcga aaagacaatc agcaaggcca agggccagcc acgggagccc
     5641 caggtgtaca ccctgccccc cagccgggag gagatgacca agaaccaggt gtccctgacc
     5701 tgtctggtga agggcttcta ccccagcgat atcgccgtgg agtgggagag caacggccag
     5761 cccgagaaca actacaagac caccccccca gtgctggaca gcgacggcag cttcttcctg
     5821 tacagcaagc tgaccgtgga caagtccagg tggcagcagg gcaacgtgtt cagctgcagc
     5881 gtgatgcacg aggccctgca caaccactac acccagaagt ccctgagcct gagccccggc
     5941 ggcagcggtc ttaatgatat tttcgaggct cagaagattg agtggcatga acaccaccac
     6001 catcaccact aaggccggcc tgtagtcact ggcacaacgc gttacccggt aagccaatcg
     6061 ggtatacacg gtcgtcatac tgcagacagg gttcttctac tttgcaagat agtcttgagt
     6121 agtaaaataa atagatagag aaaatcagcc tcgactgtgc cttctagttg ccagccatct
     6181 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt
     6241 tcctaataaa atgaggaaat tgcatcacaa cactcaaccc tatctcggtc tattcttttg
     6301 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     6361 aatttaacgc gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
     6421 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     6481 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     6541 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     6601 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc
     6661 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     6721 gctcccggga gcttgtatat ccattttcgg atctgatcag cacgtgttga caattaatca
     6781 tcggcatagt atatcggcat agtataatac gacaaggtga ggaactaaac catgaccgag
     6841 tacaagccca cggtgcgcct cgccacccgc gacgacgtcc ccagggccgt acgcaccctc
     6901 gccgccgcgt tcgccgacta ccccgccacg cgccacaccg tcgatccgga ccgccacatc
     6961 gagcgggtca ccgagctgca agaactcttc ctcacgcgcg tcgggctcga catcggcaag
     7021 gtgtgggtcg cggacgacgg cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa
     7081 gcgggggcgg tgttcgccga gatcggcccg cgcatggccg agttgagcgg ttcccggctg
     7141 gccgcgcagc aacagatgga aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg
     7201 ttcctggcca ccgtcggcgt ctcgcccgac caccagggca agggtctggg cagcgccgtc
     7261 gtgctccccg gagtggaggc ggccgagcgc gccggggtgc ccgccttcct ggagacatcc
     7321 gcgccccgca acctcccctt ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag
     7381 gtgcccgaag gaccgcgcac ctggtgcatg acccgcaagc ccggtgcctg acacgtgcta
     7441 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg
     7501 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc
     7561 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     7621 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     7681 tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcattacca
     7741 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     7801 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagcgc
     7861 tgcgatgata ccgcgagaac cacgctcacc ggctccggat ttatcagcaa taaaccagcc
     7921 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     7981 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     8041 tgccatcgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     8101 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     8161 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     8221 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     8281 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     8341 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     8401 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     8461 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     8521 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     8581 atgttgaata ctcatattct tcctt
//
