Gynogenesis is an asexual mode of reproduction common to animals and plants, in which stimuli from the sperm cell trigger the development of the unfertilized egg cell into a haploid embryo. Fine mapping restricted a major maize QTL (quantitative trait locus) responsible for the aptitude of inducer lines to trigger gynogenesis to a zone containing a single gene () coding for a patatin-like phospholipase A. In all surveyed inducer lines, carries a 4-bp insertion leading to a predicted truncated protein. This frameshift mutation is responsible for haploid induction because complementation with wild-type abolishes the haploid induction capacity. Activity of the promoter is restricted to mature pollen and pollen tube. The translational NLD::citrine fusion protein likely localizes to the sperm cell plasma membrane. In roots, the truncated protein is no longer localized to the plasma membrane, contrary to the wild-type NLD protein. In conclusion, an intact pollen-specific phospholipase is required for successful sexual reproduction and its targeted disruption may allow establishing powerful haploid breeding tools in numerous crops.

The ¢rst animal polygalacturonase (PG, EC 2.1.15) encoding cDNA, Mi-pg-1, was cloned from the plant parasitic nematode Meloidogyne incognita. The enzymatic activity of MI-PG-1 was con¢rmed after heterologous expression in Escherichia coli. The presence of a predicted signal peptide on the MI-PG-1 sequence together with the speci¢c localization of the transcripts of the Mi-pg-1 gene in the oesophageal glands of infective juveniles imply that MI-PG-1 could be secreted into plant tissues. The potential role of MI-PG-1 in parasitism is discussed. ß

SummaryLow transformation efficiency and high background of non-targeted events are major constraints to gene targeting in plants. We demonstrate here applicability in maize of a system that reduces the constraint from transformation efficiency. The system requires regenerable transformants in which all of the following elements are stably integrated in the genome: (i) donor DNA with the gene of interest adjacent to sequence for repair of a defective selectable marker, (ii) sequence encoding a rare-cutting endonuclease such as I-SceI, (iii) a target locus (TL) comprising the defective selectable marker and I-SceI cleavage site. Typically, this requires additional markers for the integration of the donor and target sequences, which may be assembled through cross-pollination of separate transformants. Inducible expression of I-SceI then cleaves the TL and facilitates homologous recombination, which is assayed by selection for the repaired marker. We used bar and gfp markers to identify assembled transformants, a dexamethasone-inducible I-SceI::GR protein, and selection for recombination events that restored an intact nptII. Applying this strategy to callus permitted the selection of recombination into the TL at a frequency of 0.085% per extracted immature embryo (29% of recombinants). Our results also indicate that excision of the donor locus (DL) through the use of flanking I-SceI cleavage sites may be unnecessary, and a source of unwanted repair events at the DL. The system allows production, from each assembled transformant, of many cells that subsequently can be treated to induce gene targeting. This may facilitate gene targeting in plant species for which transformation efficiencies are otherwise limiting.

No abstract

A new cDNA, named Mi-ace-2, encoding an acetylcholinesterase, was isolated from Meloidogyne incognita. The fulllength cDNA, carrying the trans-spliced SL1 leader sequence, was 2122 bp long with an open reading frame of 2058 bp. The predicted protein shared 32.3% identity with the previously identi ed MI ACHE and 49.1% identity with the C. elegans ACE-2. The conserved motifs involved in the catalytic triad, the choline binding site and 11 aromatic residues lining the catalytic gorge were present in the MI-ACE-2 deduced protein. RT-PCR analysis showed that Mi-ace-2 is transcribed in second stage juveniles before and after hatching, in females and in males. Phylogenetic analysis showed that MI-ACE-2 and ACE-2 were clustered in a distinct group closely related to the acetylcholinesterasessecreted by animal-parasitic nematodes.