Developmental regulation in anther tissue of plants

Details Developmental regulation in anther tissue of plants Developmental regulation in anther tissue of plants

1995-08-07

2004-05-25

Fox, David T. (Department: 1638)

Organic compounds -- part of the class 532-570 series

Organic compounds

Carbohydrates or derivatives

C536S024100, C800S287000, C800S303000, C800S306000, C435S069100, C435S468000

Reexamination Certificate

active

06740748

ABSTRACT:

FIELD OF THE INVENTION
The present invention relates generally to the use of genetic engineering to induce developmental regulation in anther tissue of plants, and more particularly to induce nuclear male sterility, and to genetic sequences useful for same.
Nucleotide and amino acid sequences are referred to herein by sequence identity numbers (SEQ ID NOs) which are defined after the bibliography. A general summary of the SEQ ID NOs is provided before the examples.
Throughout this specification and the claims which follow, unless the context requires otherwise, the word “comprise”, or variations such as “comprises” or “comprising”, will be understood to imply the inclusion of a stated element or integer or group of elements or integers but not the exclusion of any other element or integer or group of elements or integers.
Anther-specific genes are those genes that are expressed exclusively in the male reproductive tissues, rather than “house-keeping” genes which are active in all plant cells. Anther-specific genes play an important role in pollen development and, hence, in the control of seed production.
Differentiation and development of the male gametophyte of angiosperms, the pollen grain, depends partly upon transcription of the haploid genome following meiosis (Mascarenhas, 1988). The study of these coordinated events at the molecular level has been considered important in order to understand the developmentally specific regulation and functions of pollen-expressed genes. In this regard, Theerakulpisut et al (1991) studied gene expression in pollen of
Brassica campestris
. By differential screening of a mature
B. campestris
pollen cDNA library, an anther-specific clone, designated Bcp1, was isolated.
In work leading up to the present invention, the inventors undertook a detailed investigation of Bcp1 expression with the aim of isolating a genomic clone and to sub-clone and characterise the 5′ upstream regulatory regions of the genomic clone. It has been surprisingly discovered that the genomic clone of Bcp1, i.e. Bgp1, is tissue and developmentally specific thereby providing a means to enable tissue and developmental regulation in plants and in particular to produce nuclear male sterile plants. It has further been discovered that the Bgp1 gene from
B. campestris
represents a family of homologous genes from a diverse range of plants. By way of shorthand notation, a genomic clone is referred to herein by the genus and/or species of the plant from which it is isolated followed by the term “Bgp1”. A cDNA clone is referred to in similar fashion except using the term “Bcp1”.
SUMMARY OF THE INVENTION
Accordingly, one aspect of the present invention contemplates a genomic DNA isolate comprising:
(i) all or part of a gene or related genetic sequence preferentially expressed in anther tissue of a plant and substantially not expressed in non-anther tissue; and
(ii) an open reading frame having a nucleotide sequence as set forth in SEQ ID NO. 1:
ATG GGT CGC CAA AAC GCT GTC GTA GTT TTT GGC CTT GTG TTC TTG CCC

ATC CTT GGC CTC GCC GCA GCT GCC TCC TCT CCG TCT CCT TCA GCG TCA

CCC TCC AAA GCT CCG GCT GCT ACC GTA ACC GAT GTC GAA GCT CCA GTG

AGC GAG GAC ACC ATT GGA ACC ACC GAT GAC GAT GCA GCT GCT TCT CCA

GGT GAT GGT GAC GTA GCT GTG GCT CGT CCT CTA GGA AGT GAC TCC TCC

TAC GCT AGT AAT GGA CCT TCA CCT TCT ACT GAT GCT GCT GAC AGC GGC

GCC CCT GCT CTT GGC GTC TCT GCG GTC TTC GTT GGT GTT GCA TCC ATC

GCC GGT TCT TTC TTG TTT CTC
or having at least 20% similarity to all or part thereof.
The deduced amino acid sequence to the open reading frame defined in SEQ ID NO. 1 is shown in SEQ ID NO. 2.
The expression “gene or related genetic sequence” is used in is broadest sense and includes any contiguous series of nucleotides constituting an open reading frame. Generally, an open reading frame comprises at least 48 contiguous nucleotides arranged into triplets without interuption by a stop codon.
A nucleotide sequence having at least 20% similarity to all or a portion of SEQ ID NO. 1 is referred to herein as a “homologous gene”. Preferably, there is at least 20% similarity to the entire SEQ ID NO. 1 sequence. Even :more preferably, there is at least 30% similarity, still more preferably at least 45% similarity, even still more preferably at least 55-60% similarity, yet even still more preferably at least 75-95% similarity to all or part of SEQ ID NO. 1. A “part” in this context is a contiguous series of at least 20 nucleotides in SEQ ID NO. 1.
Preferably, the genomic DNA isolate is a dicotyledonous plant such as tomato, corn, rice, wheat, raddish, tobacco and oil seed rapes. Particularly preferred plants are Brassica species, Arabidopsis species and Nicotiana species.
In a most preferred embodiment, the plant is
Brassica campestris
and the genomic DNA isolate has an open reading frame with a sequence as set forth in SEQ ID NO. 1. A preferred homologous gene having at least 20% nucleotide similarity to SEQ ID NO. 1 is from
Arabidopsis thaliana
comprising an open reading frame with a nucleotide sequence as set forth in SEQ ID NO. 3:
ATG GGT CGC CAA AAC ATT GTC GTC GTG GTT GCC CTC GTC TTC ATC CGG

ATC ATT GGC CTT GCC GCA GCT GCC TCC TCT CCA TCT CCT TCA GCG TCT

CCC TCC AAA GCT CCA GCT GCC TCC AAA ACC GAT CAT GTC GAG GCT CCA

GTC ACC GAT GAC CAA ATC GGA ACC ACC GAT GAC GAT GCA GCT CCT ACT

CCT GGT GAC GGT GAC GTT GCA GTG GCT GGT CCT CTA GGA AGT GAC TCC

TCG TAC GAC AAT GCC GCT ACA GGC TCT GCT GAT TCT GCC AAA AGC GGT

GCG GCA GCT CTT GGC GTC TCT GCG GTC GTC CTT GGT GTT ACA TCA TTG

CTG GTT CTT TCT TGT TAC TCA AGT TGG GCA TTG TTT TAT GAT AAG AAG

GTT ATT TTA AAC GAA GAT TAT TAT ATG
The deduced amino acid sequence of SEQ ID NO. 3 is defined in SEQ ID NO. 4.
Another aspect of the present invention provides a genomic DNA isolate as defined above and further comprising a promoter region 5′ to the open reading frame, wherein said promoter region:
(i) is capable of directing expression in taptum and/or pollen tissue; and
(ii) comprises a nucleotide sequence as set forth in,SEQ ID NO. 5:
TATCATTCCT TTAATTTCAA GGAATTATAG AACAAAAAAT GTTCTTTATA AAAATTAAGA

AGGAACAAGG GATTCATTCC TACTATTCTG TTCTTGGTCA TTATTTTCCT CTTCATTCAT

ATTGTTTCTT TAATTGTTAC CAATTAGAAC TTTAACGAAT AAATAGTTAA TTCGTATTAT

GAGATTTACA CAATTCTTAT TCACTCAATT TGGAGTTTTA AAGATTTTTT AAAAGATTTA

TGGTGGGAAC CTTCTTCTTT TCTTATTTAT CATGATGATG ATAACCTTCC CAGCAGAATT

ATTCTTAGAA CTTTTTTTCA CATTTAGGTA TCCATGCCTA AGTAAGGCTT AGTTAAAGAT

GTTTTATAAA CTTTGATCAA AATATTCATT CAATTAATTT GAGCTTCAAC TATAAATTGT

TGTATGCATT CGTTTTAGCC TGTAAGATAT CAGACATTCA CGTTTCGATA AACAAGTATA

TAAATAATAT GAATATTGTA CATTCATTTT ATTCGGTTCA TCAACCAAAA AAAATAAAAA

TAAATATTCG TATTCATCTA TGCTTTGGCA TGGTCCGTTC TTTTTTCTTG ATTGGCTCGT

TACCATTCAA AAATATATAC CTTAGCAAAC CCATTTTTAG ACATTCCAGT TGATCTACAT

TAGATTGAAC GGTATTCCTC CTACGTAGTA AGAACGTTTT CTATTTTTCT TTGTTTCAGT

CATACAACAC AACTATATAT ACACAGCAAC CCCATCTCCT CTCCAATCAT CACAATCTCT

AACGTTAAAC CCTAAGACAA ACTAAAAGAG AGCTACGTAC AAGGAGACAG AGAGAAGA
or having at least 20% similarity to all or part thereof.
Preferred promoters comprise the promoter defined in SEQ ID NO. 5 and the promoter defined in SEQ D NO. 6 which has the following nucleotide sequence:
AAAAGCGAGA AGAAGAAGTC TGGAAGATTT GAGAGCTTAA AGTGGTCGAG TCTAAAACCC

TAACTCGCTG TTGATGGCAG AATCGTAAAT CGGAATTGAT TCATGGGCCT AACAAGACGT

TTGGGCTTAT GGGTTTAAAG CCCATCTGAT ATAAGATGAA TAGAATGTTC ATGGCAATAC

TATCATAATT TGGTTCTTTA ATAAGACACT CCTTAATACG ACGACGATTT GAAGTTGAAC

GAATCTTTTC ATATTCATTC GCATGTTCAC CAATCAAAAT CTATATCTGA ACAAGTCCAT

TTTTAGGTAC TCCAGTAGAT TTACATTGGA TTGTAAGGTA ATCCTACATC TTAGTTCACG

TTTTCTATTT TTGGTCTTGT CACTAAACAC AACTATATAT ACATATCAAA CTCATCTTCG

GAAATCATCA CAATCAATAA ACCTCAAACC CTAAAATAAA TTAAACGAGT TCTACGTAAG

AAGGAGAGAG AGAAGA
Yet another aspect of the present invention relates to a genomic DNA isolate comprising:
(i) all or part of a gene or related genetic sequence preferentially expressed in anther tissue of a plant and substantially not expressed in non-anther tissue;
(ii) a promoter region capable of directing expression in ta

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