Patent application EP20178625A (EP), filed 2020-06-05
Publication number: EP3747263A1
Title: NEW PLANTS HAVING A WHITE FOLIAGE PHENOTYPE
Abstract: [0001]    Described is a method for the generation of a Euphorbia pulcherrima (Poinsettia) plant having a white foliage phenotype, comprising the steps of:
- providing a Euphorbia pulcherrima (Poinsettia) plant without a white foliage phenotype comprising in its genome at least one functional allele of a glutathione S-transferase gene EpGST encoding a protein of SEQ ID No 3 or a functional variant of said protein with at least 90% amino acid identity, the EpGST gene comprising, in the region encoding amino acids at positions 40 - 50 of the protein of SEQ ID No 3, a stretch of 12 nucleotides consisting of a threefold CTTC repeat;
- subjecting the Euphorbia pulcherrima (Poinsettia) plant of step a. to a mutagenesis treatment to produce a mutant Euphorbia pulcherrima (Poinsettia) plant;
- selecting a mutant Euphorbia pulcherrima (Poinsettia) plant of step b., wherein at least one allele of the EpGST gene comprises a CTTC deletion in the stretch of 12 nucleotides consisting of a threefold CTTC repeat as present in the genome of the Euphorbia pulcherrima (Poinsettia) plant of step a.;
- optionally repeating steps b. and c. until all alleles of the EpGST gene in the plant genome comprise the said CTTC deletion;
- selecting a Euphorbia pulcherrima (Poinsettia) plant with white foliage phenotype being homozygous for the EpGST gene comprising the said CTTC deletion, and
- optionally further propagating said Euphorbia pulcherrima (Poinsettia) plant being homozygous for the EpGST gene comprising the said CTTC deletion.
[0002]    Further a plant, plant parts, isolated DNA, a molecular marker, and the use of the DNA to produce a molecular marker are disclosed.
IPC codes: A01H   1/    04            A I
A01H   1/    06            A I
A01H   5/    02            A I
A01H   5/    12            A I
A01H   6/    38            A I
C12N  15/    82            A I
C12N   9/    10            A I
C12Q   1/  6895            A I
Claims: number: 1
text: A method for the generation of a Euphorbia pulcherrima (Poinsettia) plant having a white foliage phenotype, comprising the steps of:
number: 2
text: Method for the generation of a Euphorbia pulcherrima (Poinsettia) plant having a white foliage phenotype of claim 1, wherein the Euphorbia pulcherrima (Poinsettia) plant provided in step a. is heterozygous for the EpGST gene, comprising one functional allele as well as one dysfunctional allele comprising the CTTC deletion, said heterozygous Euphorbia pulcherrima (Poinsettia) plant without a white foliage phenotype preferably being selected by a molecular marker suitable for the detection of the CTTC deletion.
number: 3
text: Method of claim 1 or 2, wherein the EpGST gene is selected from the group comprising
number: 4
text: Method of claim 3, wherein the gene encodes
number: 5
text: Method of any of the claims 1 to 4, wherein the EpGST cDNA has a nucleotide sequence of SEQ ID No 1.
number: 6
text: Method of any of the preceding claims, wherein
number: 7
text: A white-foliaged Euphorbia pulcherrima (Poinsettia) plant, seed, plant part, plant cell, or a plant population thereof, the said plant being derived from a non-white foliaged cultivated Euphorbia pulcherrima (Poinsettia) plant, said non-white foliaged cultivated Euphorbia pulcherrima (Poinsettia) plant comprising in its genome a glutathione S-transferase gene EpGST encoding a protein of SEQ ID No 3, or encoding a functional homolog or variant of said protein with at least 60%, preferably 70% or 80%, more 90%, most preferably 95% amino acid identity with the said protein of SEQ ID No 3, the EpGST gene comprising, in the region encoding amino acids at positions 40 - 50 of the protein of SEQ ID No 3, a stretch of 12 nucleotides consisting of a threefold CTTC repeat; the white-foliaged derivative plant being obtainable by the method of any of the preceding claims, which derivative plant is genetically identical to the non-white foliaged cultivated plant, but for
number: 8
text: A white-foliaged Euphorbia pulcherrima (Poinsettia) plant, seed, plant part, plant cell, or a plant population thereof of claim 7,
number: 9
text: Molecular marker capable of identifying an EpGST gene comprising the CTTC deletion as defined in any of the preceding claims, preferably being capable of identifying the CTTC deletion within positions 128-139 of the EpGST gene of SEQ ID No 1.
number: 10
text: Use of the molecular marker of claim 10 in the selection step c. of the method of any of the claims 1 - 6.
number: 11
text: Isolated genomic DNA of the EpGST gene described by SEQ ID NO: 61 or a variant thereof having at least 80% identity to the sequence described by SEQ ID NO: 1 encoding a functional homolog or variant of the protein of SEQ ID No 3, the said homolog or variant having a first domain at positions 11 - 13 of SEQ ID No 3 being AAC, AGC or AAN, where N can be any amino acid, a second domain at positions 53 - 56 of SEQ ID No 3 being LVPA, QVPA or QPVP and a third amino acid domain at positions 65 - 68 of SEQ ID No 3 being FESR, the EpGST gene comprising, in the region encoding amino acids at positions 40 - 50 of the protein of SEQ ID No 3, a stretch of 12 nucleotides consisting of a threefold CTTC repeat, or the complementary sequence thereof, or a fragment of the gene or the complementary sequence of at least 17 contiguous nucleotides, the said isolated DNA of the EpGST gene, the complementary sequence thereof, or a fragment of the gene or the complementary sequence of at least 17 contiguous nucleotides preferably comprising a CTTC deletion as identified in any of the preceding claims, preferably within positions 128-139 of SEQ ID No 1.
number: 12
text: Isolated cDNA encoding a EpGST protein described by SEQ ID NO: 3 or encoding a functional homolog or variant of said protein with at least 60%, preferably 70% or 80%, more 90%, most preferably 95% amino acid identity, the said homolog or variant having a first domain at positions 11 - 13 of SEQ ID No 3 being AAC, AGC or AAN, where N can be any amino acid, a second domain at positions 53 - 56 of SEQ ID No 3 being LVPA, QVPA or QPVP and a third amino acid domain at positions 65 - 68 of SEQ ID No 3 being FESR, or a fragment of at least 20 continuous nucleotides thereof, or the complements thereof.
number: 13
text: A method of use of the isolated DNA selected from the group consisting of (i) the sequences of any of claim 11 or 12 and (ii) the sequences of SEQ ID NO: 44 to 49, or variants with at least 95% identity therewith for the preparation of a molecular marker of claim 9 or for a method for targeted mutagenesis of a GST gene in a target plant, preferably using a continuous stretch of at least 17 nucleotides from any said isolated DNA sequences to (a) produce a guide RNA or an expression construct therefor for a CRISPR/Cas-based method of gene editing or (b) produce a silencing RNA or an expression construct therefor for a method of RNA-mediated gene silencing.
number: 14
text: A method of for the production of plants having a reduced level of anthocyanins comprising the steps of:
number: 15
text: Method for the production of plants having a reduced level of anthocyanins of claim 14, wherein:
Office action: search_report, dated 2020-12-09
Rejection reasons: novelty_epc_54
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Final disposition: pending (2020-12-09)