LOCUS       sequence-298418-        7237 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid for his-tagged sACE2 high affinity 
            variant 2.4.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7237)
  AUTHORS   Chan KK, Dorosky D, Sharma P, Abbasi SA, Dye JM, Kranz DM, Herbert 
            AS, Procko E
  TITLE     Engineering human ACE2 to optimize binding to the spike protein of 
            SARS coronavirus 2.
  JOURNAL   Science. 2020 Sep 4;369(6508):1261-1265. doi: 
            10.1126/science.abc0870. Epub 2020 Aug 4.
  PUBMED    32753553
REFERENCE   2  (bases 1 to 7237)
  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: 
            "10.1126/science.abc0870"; journalName: "Science"; date: "2020-09-4-
            4"; volume: "369"; issue: "6508"; pages: "1261-1265"
COMMENT     Sequence Label: pcDNA3-sACE2v2.4-8his
FEATURES             Location/Qualifiers
     source          1..7237
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2527..2550
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     primer_bind     complement(2594..2611)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2600..2824
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2870..3298
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2957..2976)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3167..3188
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3307..3327)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3312..3641
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3492..3627
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3554..3573
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3708..4502
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3762..3781)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4372..4391
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4676..4797
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4713..4732)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4767..4786
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4846..4862)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4846..4862)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4859..4881)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4870..4886
                     /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)."
     promoter        complement(join(4894..4900,4901..4918,4919..4924))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4939..4960
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5077..5094)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5248..5836)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5328..5347)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6007..6798,6799..6867))
                     /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     6630..6649
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6868..6972)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7047..7066)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg
      661 gctagcgcca ccatgtcaag ctcttcctgg ctccttctca gccttgttgc tgtaactgct
      721 gctcagtcca ccattgagga acaggccaag tactttttgg acaagtttaa ccacgaagcc
      781 gaagacctgt tctatcaaag ttcacttgct tcttggaatt ataacaccaa tattactgaa
      841 gagaatgtcc aaaacatgaa taatgctggg gacaaatggt ctgccttttt aaaggaacag
      901 tccacaactg cccaaatgta tccactacaa gaaattcaga atctcacagt caagcttcag
      961 ctgcaggctc ttcagcaaaa tgggtcttca gtgctctcag aagacaagag caaacggttg
     1021 aacacaattc taaatacaat gagcaccatc tacagtactg gaaaagtttg taacccagat
     1081 aatccacaag aatgcttatt acttgaacca ggtttgaatg aaataatggc aaacagttta
     1141 gactacaatg agaggctctg ggcttgggaa agctggagat ctgaggtcgg caagcagctg
     1201 aggccattat atgaagagta tgtggtcttg aaaaatgaga tggcaagagc aaatcattat
     1261 gaggactatg gggattattg gagaggagac tatgaagtaa atggggtaga tggctatgac
     1321 tacagccgcg gccagttgat tgaagatgtg gaacatacct ttgaagagat taaaccatta
     1381 tatgaacatc ttcatgccta tgtgagggca aagttgatga atgcctatcc ttcctatatc
     1441 agtccaattg gatgcctccc tgctcatttg cttggtgata tgtggggtag attttggaca
     1501 aatctgtact ctttgacagt tccctttgga cagaaaccaa acatagatgt tactgatgca
     1561 atggtggacc aggcctggga tgcacagaga atattcaagg aggccgagaa gttctttgta
     1621 tctgttggtc ttcctaatat gactcaagga ttctgggaat attccatgct aacggaccca
     1681 ggaaatgttc agaaagcagt ctgccatccc acagcttggg acctggggaa gggcgacttc
     1741 aggattctta tgtgcacaaa ggtgacaatg gacgacttcc tgacagctca tcatgagatg
     1801 gggcatatcc agtatgatat ggcatatgct gcacaacctt ttctgctaag aaatggagct
     1861 aatgaaggat tccatgaagc tgttggggaa atcatgtcac tttctgcagc cacacctaag
     1921 catttaaaat ccattggtct tctgtcaccc gattttcaag aagacaatga aacagaaata
     1981 aacttcctgc tcaaacaagc actcacgatt gttgggactc tgccatttac ttacatgtta
     2041 gagaagtgga ggtggatggt ctttaaaggg gaaattccca aagaccagtg gatgaaaaag
     2101 tggtgggaga tgaagcgaga gatagttggg gtggtggaac ctgtgcccca tgatgaaaca
     2161 tactgtgacc ccgcatctct gttccatgtt tctaatgatt actcattcat tcgatattac
     2221 acaaggaccc tttaccaatt ccagtttcaa gaagcacttt gtcaagcagc taaacatgaa
     2281 ggccctctgc acaaatgtga catctcaaac tctacagaag ctggacagaa actgttcaat
     2341 atgctgaggc ttggaaaatc agaaccctgg accctagcat tggaaaatgt tgtaggagca
     2401 aagaacatga atgtaaggcc actgctcaac tactttgagc ccttatttac ctggctgaaa
     2461 gaccagaaca agaattcttt tgtgggatgg agtaccgact ggagtccata tgcagacgga
     2521 tccggtcacc atcaccacca ccatcatcat tgataactcg agtctagagg gcccgtttaa
     2581 acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc
     2641 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag
     2701 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag
     2761 gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct
     2821 atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt
     2881 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     2941 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     3001 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     3061 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     3121 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     3181 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     3241 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa
     3301 ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     3361 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     3421 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     3481 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     3541 gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga
     3601 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta
     3661 tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag
     3721 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg
     3781 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc
     3841 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag
     3901 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
     3961 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
     4021 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
     4081 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
     4141 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
     4201 tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg
     4261 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
     4321 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
     4381 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
     4441 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
     4501 gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga
     4561 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc
     4621 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt
     4681 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     4741 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
     4801 tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc
     4861 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     4921 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     4981 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     5041 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     5101 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     5161 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     5221 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     5281 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     5341 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     5401 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     5461 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     5521 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     5581 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     5641 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     5701 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     5761 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     5821 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     5881 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     5941 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     6001 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     6061 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     6121 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     6181 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     6241 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     6301 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     6361 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     6421 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     6481 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     6541 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     6601 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     6661 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     6721 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     6781 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     6841 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     6901 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     6961 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat cgggagatct
     7021 cccgatcccc tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
     7081 tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa aatttaagct
     7141 acaacaaggc aaggcttgac cgacaattgc atgaagaatc tgcttagggt taggcgtttt
     7201 gcgctgcttc gcgatgtacg ggccagatat acgcgtt
//
