LOCUS       sequence-303121-        8908 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 8908)
  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 8908)
  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-CD101-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..8908
                     /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             6250..6294
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6295..6312
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6452..6469)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6458..6569
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6687..6707)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6692..7021
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6872..7007
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6934..6953
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7069..7116
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7135..7734
                     /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(7135..7154)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7631..7651
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    7864..7985
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7901..7920)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7955..7974
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(8038..8829,8830..8898))
                     /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     8661..8680
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(8899..8908,1..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 gagagaagtg
     2761 acagttcaga agggccctct gttcagagcc gagggctatc ctgtgtccat cggctgtaat
     2821 gtgacaggcc accagggacc tagcgagcag catttccagt ggtccgtgta cctgcctaca
     2881 aaccccacac aagaggtgca gatcatcagc accaaggacg ccgcctttag ctacgccgtg
     2941 tatactcaga gagtccgctc tggcgacgtg tacgtggaaa gagtgcaggg caatagcgtg
     3001 ctgctgcaca tcagcaagct gcagatgaag gatgccggcg agtacgagtg tcacacccct
     3061 aacaccgacg agaagtacta cggcagctac agcgccaaga ccaacctgat cgtgatcccc
     3121 gatacactga gcgccaccat gtctagccag acactgggaa aagaggaagg cgagcctctg
     3181 gctctgacct gtgaagcctc taaagccacc gctcagcaca cccacctgag cgtgacatgg
     3241 tatctgactc aggatggcgg cggaagccag gccacagaga tcattagcct gagcaaggac
     3301 ttcatcctgg tgcctggacc tctgtacacc gagagatttg ccgcttccga cgtgcagctg
     3361 aacaagctgg gccctaccac cttcagactg agcatcgaaa gactgcagag cagcgaccag
     3421 ggccagctgt tttgtgaagc caccgagtgg attcaggacc ccgacgagac atggatgttc
     3481 atcaccaaga agcagaccga ccagaccaca ctgagaatcc agcctgccgt gaaggacttc
     3541 caagtgaaca tcaccgccga cagcctgttt gccgagggaa aacctctgga actcgtgtgc
     3601 ctggtggtgt cctctggaag agatcctcag ctgcaaggca tctggttctt caacggaacc
     3661 gagatcgccc acatcgatgc tggcggagtg ctgggcctga agaacgacta caaagagaga
     3721 gccagccagg gcgagctgca agtgtctaaa ctgggaccta aggccttcag cctgaagatt
     3781 ttcagtctgg gccctgagga cgagggcgcc tatagatgtg tggtggccga agtgatgaag
     3841 accagaacag gctcttggca ggtcctgcag agaaagcaga gccccgatag ccacgtgcac
     3901 ctgagaaaac ctgccgccag atctgtcgtg atgagcacca agaacaagca gcaggtcgtg
     3961 tgggaaggcg aaaccctggc ctttctgtgt aaagccggcg gagctgagag ccctttgtct
     4021 gtgtcttggt ggcacatccc cagagatcag acccagcctg agtttgtggc cggcatggga
     4081 caagatggca ttgtgcagct gggagccagc tacggcgtgc catcttatca cggcaacacc
     4141 cggctggaaa agatggactg ggccaccttt cagctggaaa tcaccttcac cgccatcacc
     4201 gatagcggca cctacgagtg cagagtgtcc gagaagtcca gaaatcaggc ccgggatctg
     4261 agctggaccc agaaaatcag cgtgaccgtg aagtccctgg aaagcagcct gcaggtttcc
     4321 ctgatgagca gacagcccca agtgatgctg accaacacct tcgacctgag ctgtgtcgtc
     4381 agagccggct attctgacct gaaggtgcca ctgaccgtga cctggcagtt tcagcctgcc
     4441 agcagccaca tcttccacca gctgatcaga atcacccaca acggcacaat cgagtggggc
     4501 aacttcctga gccggttcca gaaaaagacc aaggtgtcac agtccctgtt tagaagccag
     4561 ctgctggtgc acgacgccac agaggaagaa acaggcgtct accagtgcga ggtggaagtg
     4621 tacgaccgga acagcctgta caacaacaga cctcctagag ccagcgccat ctctcaccct
     4681 ctgagaattg ccgtgacact gcccgagagc aagctgaaag tgaacagcag atcccaggtg
     4741 caagagctga gcatcaacag caacaccgac atcgagtgca gcatcctgag caggtccaat
     4801 ggcaacctgc agctggccat catctggtac tttagccccg tgtccaccaa cgcctcttgg
     4861 ctgaagatcc tggaaatgga ccagacaaac gtgatcaaga ccggcgacga gtttcacaca
     4921 ccccagcgga agcagaagtt ccacacagag aaggtgtccc aggacctgtt ccagctgcat
     4981 atcctgaacg tggaagatag cgaccggggc aagtaccact gtgccgtgga agaatggctc
     5041 ctgagcacaa acggcacctg gcacaaactg ggcgagaaga agtctggcct gaccgagctg
     5101 aagctgaagc ctacaggctc taaagtgcgg gtgtccaagg tgtactggac cgagaatgtg
     5161 accgagcata gagaagtggc catccggtgt agcctggaat ctgtgggaag cagcgccaca
     5221 ctgtacagcg tgatgtggta ctggaacaga gagaacagcg gcagcaagct cctggtgcat
     5281 ctgcaacacg atggcctgct ggaatatggc gaggaaggac tgcggcggca tctgcactgt
     5341 tacagaagca gctccaccga cttcgtcctg aagctgcacc aggtcgagat ggaagatgcc
     5401 ggaatgtact ggtgtagagt ggccgagtgg cagctgcatg gacaccctag caaatggatc
     5461 aaccaggcca gcgacgagtc ccagagaatg gtgctgacag tgctgccaag cgagcccaca
     5521 ctgcctagca gaatctgtag ctctgcccct ggcgcgccgg gcggcgacaa gacccacacc
     5581 tgccccccct gcccagctcc agaactgctg ggagggcctt ccgtgttcct gttccccccc
     5641 aagcccaagg acaccctgat gatcagcagg acccccgagg tgacctgcgt ggtggtggcc
     5701 gtgtcccacg aggacccaga ggtgaagttc aactggtacg tggacggcgt ggaggtgcac
     5761 aacgccaaga ccaagcccag agaggagcag tacaacagca cctacagggt ggtgtccgtg
     5821 ctgaccgtgc tgcaccagga ctggctgaac ggcaaagaat acaagtgcaa agtctccaac
     5881 aaggccctgg ctgccccaat cgaaaagaca atcagcaagg ccaagggcca gccacgggag
     5941 ccccaggtgt acaccctgcc ccccagccgg gaggagatga ccaagaacca ggtgtccctg
     6001 acctgtctgg tgaagggctt ctaccccagc gatatcgccg tggagtggga gagcaacggc
     6061 cagcccgaga acaactacaa gaccaccccc ccagtgctgg acagcgacgg cagcttcttc
     6121 ctgtacagca agctgaccgt ggacaagtcc aggtggcagc agggcaacgt gttcagctgc
     6181 agcgtgatgc acgaggccct gcacaaccac tacacccaga agtccctgag cctgagcccc
     6241 ggcggcagcg gtcttaatga tattttcgag gctcagaaga ttgagtggca tgaacaccac
     6301 caccatcacc actaaggccg gcctgtagtc actggcacaa cgcgttaccc ggtaagccaa
     6361 tcgggtatac acggtcgtca tactgcagac agggttcttc tactttgcaa gatagtcttg
     6421 agtagtaaaa taaatagata gagaaaatca gcctcgactg tgccttctag ttgccagcca
     6481 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
     6541 ctttcctaat aaaatgagga aattgcatca caacactcaa ccctatctcg gtctattctt
     6601 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     6661 aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     6721 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     6781 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     6841 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     6901 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     6961 tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     7021 aaagctcccg ggagcttgta tatccatttt cggatctgat cagcacgtgt tgacaattaa
     7081 tcatcggcat agtatatcgg catagtataa tacgacaagg tgaggaacta aaccatgacc
     7141 gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg tccccagggc cgtacgcacc
     7201 ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca ccgtcgatcc ggaccgccac
     7261 atcgagcggg tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct cgacatcggc
     7321 aaggtgtggg tcgcggacga cggcgccgcg gtggcggtct ggaccacgcc ggagagcgtc
     7381 gaagcggggg cggtgttcgc cgagatcggc ccgcgcatgg ccgagttgag cggttcccgg
     7441 ctggccgcgc agcaacagat ggaaggcctc ctggcgccgc accggcccaa ggagcccgcg
     7501 tggttcctgg ccaccgtcgg cgtctcgccc gaccaccagg gcaagggtct gggcagcgcc
     7561 gtcgtgctcc ccggagtgga ggcggccgag cgcgccgggg tgcccgcctt cctggagaca
     7621 tccgcgcccc gcaacctccc cttctacgag cggctcggct tcaccgtcac cgccgacgtc
     7681 gaggtgcccg aaggaccgcg cacctggtgc atgacccgca agcccggtgc ctgacacgtg
     7741 ctacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc
     7801 cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac
     7861 cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     7921 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     7981 tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatta
     8041 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     8101 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     8161 cgctgcgatg ataccgcgag aaccacgctc accggctccg gatttatcag caataaacca
     8221 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     8281 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     8341 tgttgccatc gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     8401 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     8461 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     8521 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     8581 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     8641 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     8701 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     8761 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     8821 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     8881 gaaatgttga atactcatat tcttcctt
//
