LOCUS       sequence-420997-        7198 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 7198)
  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 7198)
  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-AMIGO3-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..7198
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     453..472
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(691..782)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     824..843
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     849..868
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      954..1542
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1443..1462
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1696..1713
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     1811..1828
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1812..1828
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    2284..2300
                     /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        2467..2846
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2847..3050
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3001..3021
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     3047..3071
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        3281..3299
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     3281..3298
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     3312..3331
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             5240..5284
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             5285..5302
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(5442..5459)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5448..5559
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5677..5697)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5682..6011
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5862..5997
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        6059..6106
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6125..6724
                     /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(6125..6144)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6621..6641
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6854..6975
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6891..6910)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6945..6964
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(7028..7198,1..621,622..690))
                     /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"
ORIGIN
        1 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
       61 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
      121 tttgcgcaac gttgttgcca tcgctacagg catcgtggtg tcacgctcgt cgtttggtat
      181 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
      241 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
      301 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
      361 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
      421 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
      481 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
      541 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
      601 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
      661 aagggcgaca cggaaatgtt gaatactcat attcttcctt tttcaatatt attgaagcat
      721 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
      781 aataggggtc agtgttacaa ccaattaacc aattctgaac attatcgcga gcccatttat
      841 acctgaatat ggctcataac accccttgct catgaccaaa atcccttaac gtgagttacg
      901 cgcgcgtcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
      961 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     1021 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     1081 gcgcagatac caaatactgt tcttctagtg tagccgtagt tagcccacca cttcaagaac
     1141 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     1201 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     1261 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     1321 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     1381 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     1441 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     1501 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     1561 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     1621 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     1681 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aaggcgagag tagggaactg
     1741 ccaggcatca aactaagcag aaggcccctg acggatggcc tttttgcgtt tctacaaact
     1801 ctttctgtgt tgtaaaacga cggccagtct taagctcggg ccccctgggc ggttctgata
     1861 acgagtaatc gttaatccgc aaataacgta aaaacccgct tcggcgggtt tttttatggg
     1921 gggagtttag ggaaagagca tttgtcagaa tatttaaggg cgcctgtcac tttgcttgat
     1981 atatgagaat tatttaacct tataaatgag aaaaaagcaa cgcactttaa ataagatacg
     2041 ttgctttttc gattgatgaa cacctataat taaactattc atctattatt tatgattttt
     2101 tgtatataca atatttctag tttgttaaag agaattaaga aaataaatct cgaaaataat
     2161 aaagggaaaa tcagtttttg atatcaaaat tatacatgtc aacgataata caaaatataa
     2221 tacaaactat aagatgttat cagtatttat tatcatttag aataaatttt gtgtcgccct
     2281 taattgtgag cggataacaa ttacgagctt catgcacagt ggcgttgact caatattggc
     2341 cattagccat attattcatt ggttatatag cataaatcaa tattggctat tggccattgc
     2401 atacgttgta tctatatcat aatatgtaca tttatattgg ctcatgtcca atatgaccgc
     2461 catgttggca ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc
     2521 atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac
     2581 cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa
     2641 tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag
     2701 tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc
     2761 ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct
     2821 acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg
     2881 gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt
     2941 tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga
     3001 cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct cgtttagtga
     3061 accgtcagat cgcctggaga cgccatccac gctgttttga cctccataga agacaccggg
     3121 accgatccag cctccgcggc cgggaacggt gcattggaac gcggattccc cgtgccaaga
     3181 gtgacgtaag taccgcctat agagtctata ggcccacccc cttggcttcg ttagaacgcg
     3241 gctacaatta atacataacc ttatgtatca tacacatacg atttaggtga cactatagaa
     3301 taacatccac tttgcctttc tctccacagg tgtccactcc caggtccaac tgcacctcgg
     3361 ttctatcgaa aacgcgcctg ctagcgccac catgaagacc atcattgccc tgagctacat
     3421 cttctgcctg gtgttcgctg cggccgccac acctgattct gagggctttc cacctcgggc
     3481 tctgcacaac tgcccctaca agtgtatctg tgccgccgac ctgctgagct gtacaggact
     3541 gggactgcag gatgtgcctg ccgaacttcc tgccgctaca gccgatctgg acctgtctca
     3601 taacgccctg cagagactga ggcctggatg gctggctcct ctgtttcagc tgagagccct
     3661 gcacctggac cacaacgaac tggatgctct cggcagaggc gtgttcgtga atgcctctgg
     3721 actgagactg ctggacctga gcagcaacac cctgagagca ctgggcagac acgatctgga
     3781 tggactggga gccctggaaa agctgctgct gttcaacaac cggctggtgc atctggacga
     3841 gcacgccttt catggactga gggctctgag ccacctgtac ctgggctgta atgagctggc
     3901 ctccttcagc ttcgaccatc tgcacggact gagcgccaca catctgctga cactggatct
     3961 gtccagcaac agactgggcc acatcagcgt gccagaactg gcagctctgc ctgccttcct
     4021 gaagaacggc ctgtatctgc ataacaaccc tctgccttgc gactgccggc tgtaccatct
     4081 gctgcaaaga tggcatcaga ggggcctgtc tgccgtgcga gatttcgcca gagaatacgt
     4141 gtgcctggcc ttcaaggtgc ccgccagtag agtgcggttc ttccagcaca gccgggtgtt
     4201 cgagaattgc tctagcgccc ctgctctggg cctcgaaaga ccagaggaac acctgtatgc
     4261 cctcgtgggc agaagcctga gactgtactg caatacctcc gtgcctgcca tgagaatcgc
     4321 ctgggtttcc ccacagcaag agctgcttag agcccctgga agcagagatg gctctattgc
     4381 cgtgctggcc gatggctctc tggccatcgg aaatgtgcaa gagcagcacg ccggcctgtt
     4441 tgtgtgtctg gcaacaggac ctcggctgca ccacaatcag acccacgagt acaacgtgtc
     4501 cgtgcacttc cccagacctg agcctgaggc cttcaacaca ggcgcgccgg gcggcgacaa
     4561 gacccacacc tgccccccct gcccagctcc agaactgctg ggagggcctt ccgtgttcct
     4621 gttccccccc aagcccaagg acaccctgat gatcagcagg acccccgagg tgacctgcgt
     4681 ggtggtggcc gtgtcccacg aggacccaga ggtgaagttc aactggtacg tggacggcgt
     4741 ggaggtgcac aacgccaaga ccaagcccag agaggagcag tacaacagca cctacagggt
     4801 ggtgtccgtg ctgaccgtgc tgcaccagga ctggctgaac ggcaaagaat acaagtgcaa
     4861 agtctccaac aaggccctgg ctgccccaat cgaaaagaca atcagcaagg ccaagggcca
     4921 gccacgggag ccccaggtgt acaccctgcc ccccagccgg gaggagatga ccaagaacca
     4981 ggtgtccctg acctgtctgg tgaagggctt ctaccccagc gatatcgccg tggagtggga
     5041 gagcaacggc cagcccgaga acaactacaa gaccaccccc ccagtgctgg acagcgacgg
     5101 cagcttcttc ctgtacagca agctgaccgt ggacaagtcc aggtggcagc agggcaacgt
     5161 gttcagctgc agcgtgatgc acgaggccct gcacaaccac tacacccaga agtccctgag
     5221 cctgagcccc ggcggcagcg gtcttaatga tattttcgag gctcagaaga ttgagtggca
     5281 tgaacaccac caccatcacc actaaggccg gcctgtagtc actggcacaa cgcgttaccc
     5341 ggtaagccaa tcgggtatac acggtcgtca tactgcagac agggttcttc tactttgcaa
     5401 gatagtcttg agtagtaaaa taaatagata gagaaaatca gcctcgactg tgccttctag
     5461 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     5521 tcccactgtc ctttcctaat aaaatgagga aattgcatca caacactcaa ccctatctcg
     5581 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     5641 ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg
     5701 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     5761 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     5821 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     5881 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     5941 aggccgcctc tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     6001 gcttttgcaa aaagctcccg ggagcttgta tatccatttt cggatctgat cagcacgtgt
     6061 tgacaattaa tcatcggcat agtatatcgg catagtataa tacgacaagg tgaggaacta
     6121 aaccatgacc gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg tccccagggc
     6181 cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca ccgtcgatcc
     6241 ggaccgccac atcgagcggg tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct
     6301 cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg gtggcggtct ggaccacgcc
     6361 ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc ccgcgcatgg ccgagttgag
     6421 cggttcccgg ctggccgcgc agcaacagat ggaaggcctc ctggcgccgc accggcccaa
     6481 ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc gaccaccagg gcaagggtct
     6541 gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag cgcgccgggg tgcccgcctt
     6601 cctggagaca tccgcgcccc gcaacctccc cttctacgag cggctcggct tcaccgtcac
     6661 cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc atgacccgca agcccggtgc
     6721 ctgacacgtg ctacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg
     6781 aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt
     6841 cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     6901 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     6961 catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc
     7021 atggtcatta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     7081 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     7141 ctggccccag cgctgcgatg ataccgcgag aaccacgctc accggctccg gatttatc
//
