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公开(公告)号：US20210340532A1
公开(公告)日：2021-11-04
申请号：US16469677
申请日：2017-08-16
发明人： John Van Der Oost , Richard Van Kranenburg , Elleke Fenna Bosma , Ioannis Mougiakos , Prarthana Mohanraju
IPC分类号： C12N15/113 , C12N9/22 , C12N15/90 , C12N15/10
摘要： Thermostable Cas9 nucleases. The present invention relates to the field of genetic engineering and more particularly to nucleic acid editing and genome modification. The present invention provides an isolated Cas protein or polypeptide fragment thereof having an amino acid sequence of SEQ ID NO: 1 or a sequence of at least 77% identity therewith. The Cas protein or polypeptide is capable of binding, cleaving, marking or modifying a double stranded target polynucleotide at a temperature in the range 20° C. and 100° C. inclusive. The invention further provides isolated nucleic acid molecules encoding said Cas9 nucleases, expression vectors and host cells. The invention also provides PAM sequences recognized by the Cas protein or polypeptide, The Cas9 nucleases disclosed herein provide novel tools for genetic engineering in general, in particular at elevated temperatures and are of particular value in the genetic manipulation of thermophilic organisms; particularly microorganisms.
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公开(公告)号：US11976306B2
公开(公告)日：2024-05-07
申请号：US16469683
申请日：2017-08-16
发明人： John Van Der Oost , Richard Van Kranenburg , Elleke Fenna Bosma , Ioannis Mougiakos , Prarthana Mohanraju
IPC分类号： C12N9/22 , C12N15/10 , C12N15/11 , C12N15/113 , C12N15/90
CPC分类号： C12N9/22 , C12N15/102 , C12N15/11 , C12N15/113 , C12N15/902 , C12N15/907 , C12N2310/20 , C12N2330/51
摘要： Thermostable Cas9 nucleases. The present invention relates to the field of genetic engineering and more particularly to nucleic acid editing and genome modification. The present invention provides an isolated Cas protein or polypeptide fragment thereof having an amino acid sequence of SEQ ID NO: 1 or a sequence of at least 77% identity therewith. The Cas protein or polypeptide is capable of binding, cleaving, marking or modifying a double stranded target polynucleotide at a temperature in the range 20° C. and 100° C. inclusive. The invention further provides isolated nucleic acid molecules encoding said Cas9 nucleases, expression vectors and host cells. The invention also provides PAM sequences recognized by the Cas protein or polypeptide, The Cas9 nucleases disclosed herein provide novel tools for genetic engineering in general, in particular at elevated temperatures.
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公开(公告)号：US12071639B2
公开(公告)日：2024-08-27
申请号：US16469691
申请日：2017-12-14
发明人： John Van Der Oost , Richard Van Kranenburg , Elleke Fenna Bosma , Ioannis Mougiakos , Prarthana Mohanraju
IPC分类号： C12N9/22 , C12N15/10 , C12N15/11 , C12N15/113 , C12N15/90
CPC分类号： C12N9/22 , C12N15/102 , C12N15/11 , C12N15/113 , C12N15/902 , C12N15/907 , C12N2310/20 , C12N2330/51
摘要： A polynucleotide encoding a ThermoCas9 protein from Geobacillus thermodenitrificans and a constitutive promoter are used to engineer eukaryotic cells, e.g. fungi, yeast or algae, so that the ThermoCas9 endonuclease is integrated and expressed from the genome of the cell. Then, a second expression plasmid is used to transfect these ThermoCas9 expressing cells, the second plasmid containing an inducible promoter and a polynucleotide encoding a guide RNA. The guide RNA combines with the ThermoCas9 to provide the targeted endonuclease activity to cleave the cell DNA at a desired locus or gene of interest. A repair-oligo is also provided to the cell whereby following DNA cleavage, homologous recombination takes place in the cell with the repair-oligo so that either a deletion or substitution of nucleotides in the locus or gene of interest is achieved. Expression vectors and methods of using the vectors to achieve ThermoCas9 mediated gene editing are described whereby higher temperatures, e.g. greater than 30° C., are used.
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公开(公告)号：US11939605B2
公开(公告)日：2024-03-26
申请号：US17575952
申请日：2022-01-14
发明人： John Van Der Oost , Richard Van Kranenburg , Elleke Fenna Bosma , Ioannis Mougiakos , Prarthana Mohanraju
CPC分类号： C12N9/22 , C12N15/102 , C12N15/11 , C12N15/113 , C12N15/902 , C12N15/907 , C12N2310/20 , C12N2330/51
摘要： Thermostable Cas9 nucleases. The present invention relates to the field of genetic engineering and more particularly to nucleic acid editing and genome modification. The present invention provides an isolated Cas protein or polypeptide fragment thereof having an amino acid sequence of SEQ ID NO: 1 or a sequence of at least 77% identity therewith. The Cas protein or polypeptide is capable of binding, cleaving, marking or modifying a double stranded target polynucleotide at a temperature in the range 20° C. and 100° C. inclusive. The invention further provides isolated nucleic acid molecules encoding the Cas9 nucleases, expression vectors and host cells. The invention also provides PAM sequences recognized by the Cas protein or polypeptide, The Cas9 nucleases disclosed herein provide novel tools for genetic engineering in general, in particular at elevated temperatures and are of particular value in the genetic manipulation of thermophilic organisms; particularly microorganisms.
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公开(公告)号：US20220213455A1
公开(公告)日：2022-07-07
申请号：US17575952
申请日：2022-01-14
发明人： John Van Der Oost , Richard Van Kranenburg , Elleke Fenna Bosma , Ioannis Mougiakos , Prarthana Mohanraju
IPC分类号： C12N9/22 , C12N15/11 , C12N15/90 , C12N15/10 , C12N15/113
摘要： Thermostable Cas9 nucleases. The present invention relates to the field of genetic engineering and more particularly to nucleic acid editing and genome modification. The present invention provides an isolated Cas protein or polypeptide fragment thereof having an amino acid sequence of SEQ ID NO: 1 or a sequence of at least 77% identity therewith. The Cas protein or polypeptide is capable of binding, cleaving, marking or modifying a double stranded target polynucleotide at a temperature in the range 20° C. and 100° C. inclusive. The invention further provides isolated nucleic acid molecules encoding said Cas9 nucleases, expression vectors and host cells. The invention also provides PAM sequences recognized by the Cas protein or polypeptide, The Cas9 nucleases disclosed herein provide novel tools for genetic engineering in general, in particular at elevated temperatures and are of particular value in the genetic manipulation of thermophilic organisms; particularly microorganisms.
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公开(公告)号：US11326162B2
公开(公告)日：2022-05-10
申请号：US16469677
申请日：2017-08-16
发明人： John Van Der Oost , Richard Van Kranenburg , Elleke Fenna Bosma , Ioannis Mougiakos , Prarthana Mohanraju
IPC分类号： C12N15/113 , C12N9/22 , C12N15/90 , C12N15/10
摘要： Thermostable Cas9 nucleases. The present invention relates to the field of genetic engineering and more particularly to nucleic acid editing and genome modification. The present invention provides an isolated Cas protein or polypeptide fragment thereof having an amino acid sequence of SEQ ID NO: 1 or a sequence of at least 77% identity therewith. The Cas protein or polypeptide is capable of binding, cleaving, marking or modifying a double stranded target polynucleotide at a temperature in the range 20° C. and 100° C. inclusive. The invention further provides isolated nucleic acid molecules encoding the Cas9 nucleases, expression vectors and host cells. The invention also provides PAM sequences recognized by the Cas protein or polypeptide, The Cas9 nucleases disclosed herein provide novel tools for genetic engineering in general, in particular at elevated temperatures and are of particular value in the genetic manipulation of thermophilic organisms; particularly microorganisms.
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公开(公告)号：US11242513B2
公开(公告)日：2022-02-08
申请号：US16469674
申请日：2016-12-14
IPC分类号： C12N9/22 , C12N15/90 , C12N15/11 , C12N15/113 , C12N15/63
摘要： The present invention relates to the field of genetic engineering and more particularly to nucleic acid editing and genome modification. The present invention provides an isolated Cas protein or polypeptide fragment thereof having an amino acid sequence of SEQ ID NO: 1 or a sequence of at least 77% identity therewith. The Cas protein or polypeptide is capable of binding, cleaving, marking or modifying a double stranded target polynucleotide at a temperature in the range 30° C. and 100° C. inclusive. The invention further provides isolated nucleic acid molecules encoding the Cas9 nucleases, expression vectors and host cells. The invention also provides PAM sequences recognized by the Cas protein or polypeptide, The Cas9 nucleases disclosed herein provide novel tools for genetic engineering at elevated temperatures and are of particular value in the genetic manipulation of thermophilic organisms; particularly microorganisms.
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公开(公告)号：US20190322993A1
公开(公告)日：2019-10-24
申请号：US16469683
申请日：2017-08-16
发明人： John Van Der Oost , Richard Van Kranenburg , Elleke Fenna Bosma , Ioannis Mougiakos , Prarthana Mohanraju
摘要： Thermostable Cas9 nucleases. The present invention relates to the field of genetic engineering and more particularly to nucleic acid editing and genome modification. The present invention provides an isolated Cas protein or polypeptide fragment thereof having an amino acid sequence of SEQ ID NO: 1 or a sequence of at least 77% identity therewith. The Cas protein or polypeptide is capable of binding, cleaving, marking or modifying a double stranded target polynucleotide at a temperature in the range 20° C. and 100° C. inclusive. The invention further provides isolated nucleic acid molecules encoding said Cas9 nucleases, expression vectors and host cells. The invention also provides PAM sequences recognized by the Cas protein or polypeptide, The Cas9 nucleases disclosed herein provide novel tools for genetic engineering in general, in particular at elevated temperatures.
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