Molecular markers of plant embryogenesis

Abstract

The present invention relates generally to a molecular marker for a plant physiological process and more particularly for plant embryogenesis. The molecular marker is, in one form, a genetic sequence from a monocot plant such as but not limited to oil-palm plants. In another form, the molecular marker is a polypeptide encoded by said genetic sequence. More particularly, the molecular marker of the present invention enables embryogenic tissue to be detected in vitro. The early detection of embryogenic tissue enables non-embryogenic tissue to be discarded. The ability to detect embryogenesis facilitates maximization of embryogenic potential. The present invention further contemplates a molecular marker comprising in one form a sequence of nucleotides encoding an antioxidant or in another form a sequence of amino acids defining a polypeptide having antioxidant activity. The antioxidant according to this aspect of the present invention is particularly useful in tablet or cream form as an anti-aging agent. The molecular markers of the present invention therefore also have uses in the inhibition or retardation of apoptotic processes. Such an effect has benefits in both plant and animal cells. The present invention further contemplates a promoter sequence encoding the molecular marker and its use in generating male sterile plants.

1 . An isolated nucleic acid molecule encoding a polypeptide comprising an amino acid sequence substantially as set forth in SEQ ID NO:2 or an amino acid sequence having at least about 71% similarity to SEQ ID NO:2, wherein said polypeptide is present in plant zygotic embryos or embryogenic callus and is substantially not present in non-embryogenic tissue. 
     
     
         2 . An isolated nucleic acid molecule of  claim 1 , wherein said nucleic acid molecule comprises a sequence of nucleotides substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3 or its complementary form, or a nucleotide sequence having at least about 71% similarity to SEQ ID NO:1 or SEQ ID NO:3 or its complementary form or a nucleotide sequence capable of hybridizing to SEQ ID NO:1 or SEQ ID NO:3 or its complementary form under low stringency conditions. 
     
     
         3 . An isolated nucleic acid molecule of  claim 1 , wherein the nucleic acid molecule is developmentally regulated. 
     
     
         4 . An isolated nucleic acid molecule of  claim 1 ,  2  or  3 , wherein the nucleic acid molecule is expressed substantially in embryogenic material of oil-palm plants or related plants but not in non-embryogenic material. 
     
     
         5 . An isolated nucleic acid molecule of  claim 1 , wherein the nucleic acid molecule comprises the nucleotide sequence substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3. 
     
     
         6 . An isolated nucleic acid molecule of  claim 1 , wherein the nucleic acid molecule comprises the nucleotide sequence substantially as set forth in SEQ ID NO:1. 
     
     
         7 . An isolated nucleic acid molecule of  claim 1 , wherein the nucleic acid molecule comprises the nucleotide sequence substantially as set forth in SEQ ID NO:3. 
     
     
         8 . A genetic construct comprising a nucleic acid molecule encoding a polypeptide comprising an amino acid sequence substantially as set forth in SEQ ID NO:2 or an amino acid sequence having at least about 71% similarity to SEQ ID NO:2, wherein said polypeptide is present in plant zygotic embryos or embryogenic callus and is substantially not present in non-embryogenic tissue. 
     
     
         9 . A genetic construct of  claim 8 , wherein the nucleic acid molecule is substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3 or its complementary form, or a nucleotide sequence having at least about 71% similarity to SEQ ID NO:1 or SEQ ID NO:3 or its complementary form or a nucleotide sequence capable of hybridizing to SEQ ID NO:1 or SEQ ID NO:3 or its complementary form under low stringency conditions. 
     
     
         10 . A genetic construct of  claim 8 , wherein the nucleic acid molecule is developmentally regulated. 
     
     
         11 . A genetic construct of  claim 8 ,  9  or  10 , wherein the nucleic acid molecule is expressed substantially in embryogenic material of oil-palm plants or related plants but not in non-embryogenic material. 
     
     
         12 . A genetic construct of  claim 8 , wherein the nucleic acid molecule comprises a sequence of nucleotides substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3. 
     
     
         13 . A genetic construct of  claim 12 , wherein the nucleic acid molecule comprises a sequence of nucleotides substantially as set forth in SEQ ID NO:1. 
     
     
         14 . A genetic construct of  claim 12 , wherein the nucleic acid molecule comprises a sequence of nucleotides substantially as set forth in SEQ ID NO:3. 
     
     
         15 . A genetic construct of  claim 8  or  9 , wherein said construct further comprises one or more promoter sequences or transcription termination sequences. 
     
     
         16 . A genetic construct of  claim 15 , wherein said construct further comprises one or more origins of replication and/or selectable marker gene sequences. 
     
     
         17 . A vector comprising a construct of any one of  claims 8  to  16 . 
     
     
         18 . A host cell comprising a nucleic acid molecule encoding a polypeptide comprising an amino acid sequence substantially as set forth in SEQ ID NO:2 or an amino acid sequence having at least about 71% similarity to SEQ ID NO:2, wherein said polypeptide is present in plant zygotic embryos or embryogenic callus and is substantially not present in non-embryogenic tissue. 
     
     
         19 . A host cell of  claim 18 , wherein said nucleic acid molecule comprises a sequence of nucleotides substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3 or its complementary form, or a nucleotide sequence having at least about 71% similarity to SEQ ID NO:1 or SEQ ID NO:3 or its complementary form or a nucleotide sequence capable of hybridizing to SEQ ID NO:1 or SEQ ID NO:3 or its complementary form under low stringency conditions. 
     
     
         20 . A host cell of  claim 18 , wherein the nucleic acid molecule is developmentally regulated. 
     
     
         21 . A host cell of  claim 18 ,  19  or  20  wherein the nucleic acid molecule is expressed substantially in embryogenic material of oil-palm plants or related plants but not in non-embryogenic material. 
     
     
         22 . A host cell of  claim 18 , wherein the nucleic acid molecule comprises the nucleotide sequence substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3. 
     
     
         23 . A host cell of  claim 18 , wherein the nucleic acid molecule comprises the nucleotide sequence substantially as set forth in SEQ ID NO:1. 
     
     
         24 . A host cell of  claim 18 , wherein the nucleic acid molecule comprises the nucleotide sequence substantially as set forth in SEQ ID NO:3. 
     
     
         25 . A host cell of  claim 18 , wherein the cell is a plant cell. 
     
     
         26 . A plant cell of  claim 25 , wherein the cell is from an oil-palm plant. 
     
     
         27 . An isolated polypeptide or biologically-active fragment thereof or a variant or derivative of these, said polypeptide comprising the amino acid sequence set forth in SEQ ID NO:2 or an amino acid sequence having at least about 71% similarity to SEQ ID NO:2, wherein said polypeptide is present in plant zygotic embryos or embryogenic callus and is substantially not present in non-embryogenic tissue. 
     
     
         28 . An isolated polypeptide of  claim 27 , wherein the amino acid sequence is encoded by the nucleotide sequence substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3 or a nucleotide sequence having at least about 71% similarity to SEQ ID NO:1 or SEQ ID NO:3 or its complementary form or a nucleotide sequence capable of hybridizing to SEQ ID NO:1 or SEQ ID NO:3 or its complementary form under low stringency conditions. 
     
     
         29 . An isolated polypeptide of  claim 27 , wherein the amino acid sequence is expressed substantially in embryogenic material of oil-palm plants or related plants but not in non-embryogenic material. 
     
     
         30 . An isolated polypeptide of  claim 27  or  28 , wherein the nucleotide sequence is substantially as set forth in SEQ ID NO:1. 
     
     
         31 . An isolated polypeptide of  claim 27  or  28 , wherein the nucleotide sequence is substantially as set forth in SEQ ID NO:3. 
     
     
         32 . An isolated polypeptide of any one of  claims 27  to  31 , wherein the polypeptide has antioxidant properties. 
     
     
         33 . An isolated polypeptide of  claim 32 , wherein the polypeptide is peroxiredoxin. 
     
     
         34 . A method for producing a recombinant polypeptide in a host cell or tissue, said method comprising introducing into the said cell or tissue an expression vector comprising a nucleic acid molecule wherein said nucleic acid molecule comprises a sequence of nucleotides substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3 or its complementary form, or a nucleotide sequence having at least about 71% similarity to SEQ ID NO:1 or SEQ ID NO:3 or its complementary form or a nucleotide sequence capable of hybridizing to SEQ ID NO:1 or SEQ ID NO:3 or its complementary form under low stringency conditions, wherein said nucleic acid molecule is operably linked to one or more regulatory sequences such that the nucleic acid molecule is capable of being expressed in said cell or tissue. 
     
     
         35 . A method of  claim 34 , wherein said expression vector comprises a genetic construct comprising a nucleic acid molecule comprising a sequence of nucleotides encoding an amino acid sequence substantially as set forth in SEQ ID NO:2 or an amino acid sequence having at least about 71% similarity to SEQ ID NO:2. 
     
     
         36 . A method of  claim 34 , wherein said nucleic acid molecule comprises a sequence of nucleotides substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3. 
     
     
         37 . A method of  claim 36 , wherein said nucleic acid molecule comprises a sequence of nucleotides substantially as set forth in SEQ ID NO:1. 
     
     
         38 . A method of  claim 36 , wherein said nucleic acid molecule comprises a sequence of nucleotides substantially as set forth in SEQ ID NO:3. 
     
     
         39 . A method for modulating apoptotic processes in a cell or tissue, said method comprising introducing into said cell or tissue an expression vector comprising a nucleic acid molecule, said nucleic acid molecule comprising a sequence of nucleotides substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3 or its complementary form, a nucleotide sequence having at least about 71% similarity to SEQ ID NO:1 or SEQ ID NO:3 or its complementary form or a nucleotide sequence capable of hybridizing SEQ ID NO:1 or SEQ ID NO:3 or its complementary form under low stringency conditions wherein said nucleic acid molecule is operably linked to one or more regulatory sequences such that the nucleic acid molecule is capable of being expressed in said cell or tissue. 
     
     
         40 . A method of  claim 39 , wherein said expression vector comprises a genetic construct comprising a nucleic acid molecule comprising a sequence of nucleotides encoding an amino acid sequence substantially as set forth in SEQ ID NO:2 or an amino acid sequence having at least about 71% similarity to SEQ ID NO:2. 
     
     
         41 . A method of  claim 39 , wherein said nucleic acid molecule comprises a sequence of nucleotides substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3. 
     
     
         42 . A method of  claim 41 , wherein said nucleic acid molecule comprises a sequence of nucleotides substantially as set forth in SEQ ID NO:1. 
     
     
         43 . A method of  claim 41 , wherein said nucleic acid molecule comprises a sequence of nucleotides substantially as set forth in SEQ ID NO:3. 
     
     
         44 . A method for modulating apoptotic processes in a cell, said method comprising administering to said cell an apoptotic process-controlling effective amount of a recombinant polypeptide, said polypeptide comprising the amino acid sequence set forth in SEQ ID NO:2 or an amino acid sequence having at least about 71% similarity to SEQ ID NO:2, said administration being for a time and under conditions sufficient to modulate apoptosis. 
     
     
         45 . A method for detecting embryogenic plant material, said method comprising immobilizing a sample putatively containing RNA from the material to be screened on a solid support and contacting said immobilized RNA with a labelled nucleotide sequence capable of hybridizing to all or part of an mRNA transcript corresponding to the nucleotide sequence set forth in SEQ ID NO:1 or SEQ ID NO:3 or their derivatives or homologues as defined herein and then detecting the presence of said label. 
     
     
         46 . A method of  claim 45 , wherein said nucleotide sequence is substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3. 
     
     
         47 . A method of  claim 45 , wherein said nucleotide sequence is substantially as set forth in SEQ ID NO:1. 
     
     
         48 . A method of  claim 45 , wherein said nucleotide sequence is substantially as set forth in SEQ ID NO:3. 
     
     
         49 . An antibody to a polypeptide, said polypeptide comprising a sequence of amino acids substantially as set forth in SEQ ID NO:2 or an amino acid sequence having at least 71% similarity to SEQ ID NO:2, wherein said polypeptide is present in plant zygotic embryos or embryogenic callus and is substantially not present in non-embryogenic tissue. 
     
     
         50 . An antibody of  claim 49 , wherein the amino acid sequence is encoded by a nucleotide sequence substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3 or a nucleotide sequence having at least about 71% similarity to SEQ ID NO:1 or SEQ ID NO:3 or its complementary form or a nucleotide sequence capable of hybridizing to SEQ ID NO:1 or SEQ ID NO:3 or its complementary form under low stringency conditions. 
     
     
         51 . An antibody of  claim 49 , wherein the amino acid sequence is encoded by a nucleotide sequence substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3. 
     
     
         52 . An antibody of  claim 49 , wherein the nucleotide sequence is substantially as set forth in SEQ ID NO:1. 
     
     
         53 . An antibody of  claim 49 , wherein the nucleotide sequence is substantially as set forth in SEQ ID NO:3. 
     
     
         54 . An antibody of  claim 49 , wherein the polypeptide has antioxidant properties. 
     
     
         55 . An antibody of  claim 54 , wherein the polypeptide is peroxiredoxin. 
     
     
         56 . A method for detecting a polypeptide which is indicative of the presence of embryogenic tissue in oil-palm or related plants, said method comprising contacting the tissue or an extract thereof with an antibody specific for said polypeptide or its derivatives or homologues for a time and under conditions sufficient for an antibody-polypeptide complex to form, and then detecting said complex. 
     
     
         57 . A method of  claim 56 , wherein the polypeptide comprises a sequence of amino acids substantially as set forth in SEQ ID NO:2 or an amino acid sequence having at least 71% similarity to SEQ ID NO:2. 
     
     
         58 . A method of  claim 57 , wherein the amino acid sequence is encoded by a nucleotide sequence substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3 or a nucleotide sequence having at least about 71% similarity to SEQ ID NO:1 or SEQ ID NO:3 or its complementary form or a nucleotide sequence capable of hybridizing to SEQ ID NO:1 or SEQ ID NO:3 or its complementary form under low stringency conditions. 
     
     
         59 . A method of  claim 56 , wherein the amino acid sequence is encoded by a nucleotide sequence substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3. 
     
     
         60 . A method of  claim 56 , wherein the amino acid sequence is encoded by a nucleotide sequence substantially as set forth in SEQ ID NO:1. 
     
     
         61 . A method of  claim 56 , wherein the amino acid sequence is encoded by a nucleotide sequence substantially as set forth in SEQ ID NO:3. 
     
     
         62 . A pharmaceutical composition comprising the polypeptide having an amino acid sequence as set forth in SEQ ID NO:2 or a functional homologue thereof or a molecule having at least 71% similarity to SEQ ID NO:2 and one or more pharmaceutically-acceptable carriers and/or diluents. 
     
     
         63 . A pharmaceutical composition of  claim 62 , wherein the amino acid sequence is as set forth in SEQ ID NO:2. 
     
     
         64 . A regenerated differentiated plant comprising a nucleic acid molecule encoding a polypeptide comprising an amino acid sequence substantially as set forth in SEQ ID NO:2 or an amino acid sequence having at least about 71% similarity to SEQ ID NO:2, wherein said polypeptide is present in plant zygotic embryos or embryogenic callus and is substantially not present in non-embryogenic tissue. 
     
     
         65 . A regenerated differentiated plant of  claim 64 , wherein nucleic acid molecule comprises a sequence of nucleotides substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3 or its complementary form, or a nucleotide sequence having at least about 71% similarity to SEQ ID NO:1 or SEQ ID NO:3 or its complementary form or a nucleotide sequence capable of hybridizing to SEQ ID NO:1 or SEQ ID NO:3 or its complementary form under low stringency conditions. 
     
     
         66 . A regenerated differentiated plant of  claim 64 , wherein the amino acid sequence is encoded by a nucleotide sequence substantially as set forth in SEQ ID NO:1 or SEQ ID NO:3. 
     
     
         67 . A regenerated differentiated plant of  claim 64 , wherein the amino acid sequence is encoded by a nucleotide sequence substantially as set forth in SEQ ID NO:1. 
     
     
         68 . A regenerated differentiated plant of  claim 64 , wherein the amino acid sequence is encoded by a nucleotide sequence substantially as set forth in SEQ ID NO:3. 
     
     
         69 . A regenerated differentiated plant of  claim 64 , wherein the plant is an oil palm plant.