LOCUS       sequence-466022-        9205 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 9205)
  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 9205)
  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-NPHS1-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..9205
                     /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             6547..6591
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6592..6609
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6749..6766)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6755..6866
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6984..7004)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6989..7318
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7169..7304
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7231..7250
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7366..7413
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7432..8031
                     /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(7432..7451)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7928..7948
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    8161..8282
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8198..8217)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8252..8271
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(8335..9126,9127..9195))
                     /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     8958..8977
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(9196..9205,1..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 cggccgccca acttgctatt
     2761 cctgcttccg tgcctagagg cttttgggcc ctgcctgaga atctgacagt ggtggaaggc
     2821 gccagcgtgg aactgagatg cggagtgtct acacctggat ctgccgtgca gtgggccaaa
     2881 gatggactgc tgcttggccc cgatcctaga atccctggct tccccagata cagactggaa
     2941 ggcgatcctg ccagaggcga gttccatctg cacatcgagg cctgcgatct gagcgacgat
     3001 gccgagtatg agtgccaagt gggcagatct gagatgggcc ctgagctggt gtctcccaga
     3061 gtgatcctgt ctatcctggt gcctcctaaa ctgctgctgc tgacacctga ggccggcaca
     3121 atggttacat gggtggccgg acaagagtac gtggtcaatt gtgtgtctgg cgacgccaag
     3181 cctgctcctg atatcacaat cctgctgagc ggccagacca tcagcgacat cagcgccaat
     3241 gtgaatgagg gcagccagca gaaactgttc accgtggaag ccaccgccag agtgacaccc
     3301 agaagcagcg ataacagaca gctgctcgtg tgcgaagcca gctctcctgc tctggaagcc
     3361 cctatcaagg ccagcttcac cgtgaacgtg ctgttccctc ctggacctcc tgtgattgag
     3421 tggcctggac tggatgaggg acacgttcga gctggacaga gcctggaact gccttgtgtg
     3481 gctagaggcg gaaatcctct ggctaccctg cagtggctga agaatggcca gcctgtgtct
     3541 accgcctggg gaacagaaca tacacaggcc gtggccagat ccgtgctggt catgacagtt
     3601 agacctgagg atcacggcgc ccagctgagc tgtgaagccc ataattctgt gtccgccggc
     3661 acacaagagc acggcatcac cctgcaagtg acctttccac ctagcgccat catcatcctg
     3721 ggcagcgcct ctcagaccga gaacaagaat gtgaccctgt cctgcgtgtc caagagcagc
     3781 agacctagag tgctgctgag atggtggctt ggatggcgtc agctgctgcc catggaagag
     3841 acagtgatgg atggcctgca cggcggccac atctctatga gcaatctgac cttcctggcc
     3901 agaagagagg acaacggcct gacactgaca tgcgaggcct tctccgaggc ctttaccaaa
     3961 gagacattca agaagtccct gatcctgaac gtgaagtacc ccgctcagaa gctgtggatc
     4021 gaaggccctc ctgagggaca gaaactgaga gctggaacaa gagtgcgcct cgtgtgcctg
     4081 gccattggag gaaatcctga gcctagcctg atgtggtaca aggacagcag aaccgtgacc
     4141 gagagcagac tgccccaaga gtccagaaga gtgcacctgg gctctgtgga aaagagcggc
     4201 agcaccttca gcagagaact ggtgctggtt acaggcccct ccgacaacca ggccaagttt
     4261 acctgtaaag ccggccagct gagtgcctcc acacagctgg ctgttcagtt cccacctacc
     4321 aacgtgacaa tcctggccaa cgcttctgct ctgaggcctg gggatgctct gaatctgact
     4381 tgcgtgtccg tgtccagcaa tcctccagtg aatctgagct gggacaaaga gggcgagaga
     4441 ctcgaaggtg ttgctgcccc tcctagaagg gcccctttta aaggatctgc agccgctaga
     4501 agcgtgctgc tccaggtgtc atccagagat cacggccaga gagtgacctg tagagcccat
     4561 tctgccgagc tgagagaaac cgtgtctagc ttctaccggc tcaatgtgct gtacagaccc
     4621 gagttcctgg gcgagcaggt tctggtggtt acagctgttg aacagggcga agctctgctg
     4681 cccgtgtctg tgtctgctaa tcccgctcct gaggccttca actggacctt cagaggctac
     4741 agactgagcc ctgctggcgg acccagacac aggattcttt ctagcggagc cctgcacctg
     4801 tggaacgtga ccagagccga tgatggcctg tatcagctgc actgccagaa tagcgaaggc
     4861 acagccgaag ccagactgag actggatgtg cactacgccc ctaccatcag agcactgcag
     4921 gaccccaccg aagtgaatgt cggcggatct gtggatatcg tgtgcaccgt ggacgctaac
     4981 cctatcctgc ctggcatgtt caactgggag agactgggcg aagatgagga agatcagtcc
     5041 ctggacgaca tggaaaagat cagcagaggc cctaccggca ggctgagaat tcatcatgcc
     5101 aaactggccc aggcaggcgc ctaccagtgt atcgttgata atggcgtggc ccctcctgct
     5161 cggagactgc tgagactcgt tgtcagattt gcccctcaag tggaacaccc cacacctctg
     5221 acaaaagtgg ccgctgccgg cgatagcaca tcttctgcta cactgcattg cagagccagg
     5281 ggcgtgccca acatcgtgtt tacctggacc aagaacggcg tgcccctgga cctgcaagat
     5341 cctagataca ccgagcacac ctaccaccaa ggcggagtgc atagcagcct gctgaccatt
     5401 gctaacgtgt cagccgctca ggactacgcc ctgtttacat gcacagccac aaacgccctg
     5461 ggctccgacc agacaaatat ccagctcgtg tccatcagca gacccgatcc tccatctggc
     5521 ctgaaggtgg tgtctctgac acctcactct gtgggcctcg agtggaagcc tggatttgat
     5581 ggtggactgc ctcagcggtt ctgcatcaga tatgaggccc tgggcacccc tggctttcac
     5641 tatgttgatg tggtgccacc tcaggctacc acattcaccc tgacaggcct gcagcctagc
     5701 acaagataca gagtgtggct gctggcctct aatgccctgg gagattctgg actggccgac
     5761 aagggaaccc agctgcctat tacaacaccc ggactgcacc agcctagcgg cgaacctgaa
     5821 gatcagctgc caaccgagcc accttctggc gcgccgggcg gcgacaagac ccacacctgc
     5881 cccccctgcc cagctccaga actgctggga gggccttccg tgttcctgtt cccccccaag
     5941 cccaaggaca ccctgatgat cagcaggacc cccgaggtga cctgcgtggt ggtggccgtg
     6001 tcccacgagg acccagaggt gaagttcaac tggtacgtgg acggcgtgga ggtgcacaac
     6061 gccaagacca agcccagaga ggagcagtac aacagcacct acagggtggt gtccgtgctg
     6121 accgtgctgc accaggactg gctgaacggc aaagaataca agtgcaaagt ctccaacaag
     6181 gccctggctg ccccaatcga aaagacaatc agcaaggcca agggccagcc acgggagccc
     6241 caggtgtaca ccctgccccc cagccgggag gagatgacca agaaccaggt gtccctgacc
     6301 tgtctggtga agggcttcta ccccagcgat atcgccgtgg agtgggagag caacggccag
     6361 cccgagaaca actacaagac caccccccca gtgctggaca gcgacggcag cttcttcctg
     6421 tacagcaagc tgaccgtgga caagtccagg tggcagcagg gcaacgtgtt cagctgcagc
     6481 gtgatgcacg aggccctgca caaccactac acccagaagt ccctgagcct gagccccggc
     6541 ggcagcggtc ttaatgatat tttcgaggct cagaagattg agtggcatga acaccaccac
     6601 catcaccact aaggccggcc tgtagtcact ggcacaacgc gttacccggt aagccaatcg
     6661 ggtatacacg gtcgtcatac tgcagacagg gttcttctac tttgcaagat agtcttgagt
     6721 agtaaaataa atagatagag aaaatcagcc tcgactgtgc cttctagttg ccagccatct
     6781 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt
     6841 tcctaataaa atgaggaaat tgcatcacaa cactcaaccc tatctcggtc tattcttttg
     6901 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     6961 aatttaacgc gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
     7021 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     7081 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     7141 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     7201 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc
     7261 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     7321 gctcccggga gcttgtatat ccattttcgg atctgatcag cacgtgttga caattaatca
     7381 tcggcatagt atatcggcat agtataatac gacaaggtga ggaactaaac catgaccgag
     7441 tacaagccca cggtgcgcct cgccacccgc gacgacgtcc ccagggccgt acgcaccctc
     7501 gccgccgcgt tcgccgacta ccccgccacg cgccacaccg tcgatccgga ccgccacatc
     7561 gagcgggtca ccgagctgca agaactcttc ctcacgcgcg tcgggctcga catcggcaag
     7621 gtgtgggtcg cggacgacgg cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa
     7681 gcgggggcgg tgttcgccga gatcggcccg cgcatggccg agttgagcgg ttcccggctg
     7741 gccgcgcagc aacagatgga aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg
     7801 ttcctggcca ccgtcggcgt ctcgcccgac caccagggca agggtctggg cagcgccgtc
     7861 gtgctccccg gagtggaggc ggccgagcgc gccggggtgc ccgccttcct ggagacatcc
     7921 gcgccccgca acctcccctt ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag
     7981 gtgcccgaag gaccgcgcac ctggtgcatg acccgcaagc ccggtgcctg acacgtgcta
     8041 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg
     8101 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc
     8161 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     8221 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     8281 tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcattacca
     8341 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     8401 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagcgc
     8461 tgcgatgata ccgcgagaac cacgctcacc ggctccggat ttatcagcaa taaaccagcc
     8521 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     8581 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     8641 tgccatcgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     8701 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     8761 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     8821 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     8881 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     8941 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     9001 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     9061 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     9121 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     9181 atgttgaata ctcatattct tcctt
//
