LOCUS       sequence-292588-        7210 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid for soluble ACE2 (protease domain) high
            affinity variant 4.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7210)
  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 7210)
  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-sACE2v4
FEATURES             Location/Qualifiers
     source          1..7210
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        129..508
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        509..712
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     663..683
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     757..776
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        757..775
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2695..2712)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2701..2925
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2971..3399
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3058..3077)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3268..3289
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3408..3428)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3413..3742
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3593..3728
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3655..3674
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3809..4603
                     /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(3863..3882)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4473..4492
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4777..4898
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4814..4833)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4868..4887
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4947..4963)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4947..4963)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4960..4982)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4971..4987
                     /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(4995..5001,5002..5019,5020..5025))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5040..5061
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5178..5195)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5349..5937)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5429..5448)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6108..6899,6900..6968))
                     /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     6731..6750
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6969..7073)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7148..7167)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 cgctgagtag tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg
       61 catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac gggccagata
      121 tacgcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
      181 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
      241 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
      301 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
      361 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
      421 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
      481 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg
      541 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
      601 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt
      661 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctctctggct
      721 aactagagaa cccactgctt actggcttat cgaaattaat acgactcact atagggagac
      781 ccaagctggc tagcgccacc atgtcaagct cttcctggct ccttctcagc cttgttgctg
      841 taactgctgc tcagtccacc attgaggaac aggccaagac atttttggac aagtttaacg
      901 ccgaagccga agacctgttc tatcaaagtt cacttgcttc ttggaattat aacaccaata
      961 ttactgaaga gaatgtccaa aacatgaata atgctgggga caaatggtct gcctttttaa
     1021 aggaacagtc cacaactgcc caaatgtatc cactacaaga aattcagaat ctcacagtca
     1081 agcttcagct gcaggctctt cagcaaaatg ggtcttcagt gctctcagaa gacaagagca
     1141 aacggttgaa cacaattcta aatacaatga gcaccatcta cagtactgga aaagtttgta
     1201 acccagataa tccacaagaa tgcttattac ttgaaccagg tttgaatgaa ataatggcaa
     1261 acagtttaga ctacaatgag aggctctggg cttgggaaag ctggagatct gaggtcggca
     1321 agcagctgag gccattatat gaagagtatg tggtcttgaa aaatgagatg gcaagagcaa
     1381 atcattatga ggactatggg gattattgga gaggagacta tgaagtaaat ggggtagatg
     1441 gctatgacta cagccgcggc cagttgattg aagatgtgga acataccttt gaagagatta
     1501 aaccattata tgaacatctt catgcctatg tgagggcaaa gttgatgaat gcctatcctt
     1561 cctatatcag tccaattgga tgcctccctg ctcatttgct tggtgatatg tggggtagat
     1621 tttggacaaa tctgtactct ttgacagttc cctttggaca gaaaccaaac atagatgtta
     1681 ctgatgcaat ggtggaccag gcctgggatg cacagagaat attcaaggag gccgagaagt
     1741 tctttgtatc tgttggtctt cctaatatga ctcaaggatt ctgggaatat tccatgctaa
     1801 cggacccagg aaatgttcag aaagcagtct gccatcccac agcttgggac ctggggaagg
     1861 gcgacttcag gattcttatg tgcacaaagg tgacaatgga cgacttcctg acagctcatc
     1921 atgagatggg gcatatccag tatgatatgg catatcttgc acaacctttt ctgctaagaa
     1981 atggagctaa tgaaggattc catgaagctg ttggggaaat catgtcactt tctgcagcca
     2041 cacctaagca tttaaaatcc attggtcttc tgtcacccga ttttcaagaa gacaatgaaa
     2101 cagaaataaa cttcctgctc aaacaagcac tcacgattgt tgggactctg ccatttactt
     2161 acatgttaga gaagtggagg tggatggtct ttaaagggga aattcccaaa gaccagtgga
     2221 tgaaaaagtg gtgggagatg aagcgagaga tagttggggt ggtggaacct gtgccccatg
     2281 atgaaacata ctgtgacccc gcatctctgt tccatgtttc taatgattac tcattcattc
     2341 gatattacac aaggaccctt taccaattcc agtttcaaga agcactttgt caagcagcta
     2401 aacatgaagg ccctctgcac aaatgtgaca tctcaaactc tacagaagct ggacagaaac
     2461 tgttcaatat gctgaggctt ggaaaatcag aaccctggac cctagcattg gaaaatgttg
     2521 taggagcaaa gaacatgaat gtaaggccac tgctcaacta ctttgagccc ttatttacct
     2581 ggctgaaaga ccagaacaag aattcttttg tgggatggag taccgactgg agtccatatg
     2641 cagacggatc ctgataactc gagtctagag ggcccgttta aacccgctga tcagcctcga
     2701 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     2761 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     2821 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     2881 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa
     2941 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg
     3001 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     3061 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     3121 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     3181 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     3241 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     3301 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     3361 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
     3421 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     3481 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     3541 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     3601 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     3661 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
     3721 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
     3781 tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc
     3841 tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg
     3901 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
     3961 cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc
     4021 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
     4081 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
     4141 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
     4201 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
     4261 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
     4321 cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
     4381 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
     4441 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
     4501 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
     4561 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc
     4621 gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc
     4681 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag
     4741 cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat
     4801 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     4861 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc
     4921 tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
     4981 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
     5041 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     5101 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     5161 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     5221 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     5281 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     5341 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     5401 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     5461 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     5521 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     5581 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     5641 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     5701 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     5761 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     5821 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     5881 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     5941 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     6001 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     6061 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     6121 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     6181 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     6241 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     6301 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     6361 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     6421 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     6481 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     6541 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     6601 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     6661 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     6721 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     6781 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     6841 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     6901 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     6961 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     7021 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     7081 ccccgaaaag tgccacctga cgtcgacgga tcgggagatc tcccgatccc ctatggtgca
     7141 ctctcagtac aatctgctct gatgccgcat agttaagcca gtatctgctc cctgcttgtg
     7201 tgttggaggt
//
