Phosphoenolpyruvate carboxylase (PEPC) plays a crucial role in the assimilation of CO2 during symbiotic N2 fixation in legume root nodules. In this study, an alfalfa PEPC gene (PEPC-7), whose transcripts are found at elevated levels in nodules relative to either leaves or roots, has been isolated and characterized. The intron/exon structure of this gene is identical to that of most other plant PEPC genes except for the presence of an additional intron in the 5' untranslated region. In situ RNA hybridization studies showed that PEPC transcripts were present in the nodule meristem, the infection zone, the nitrogen-fixing zone, and the parenchyma. PEPC transcripts were also found in vascular tissue of roots and nodules and in the pulvinus of petioles. In transgenic alfalfa, a chimeric reporter gene was expressed in these same regions except that little expression was found in the nodule meristem. Analyses of promoter deletions suggest that the region between -634 and -536 is of particular importance in directing transcriptional activity to the infected zone of nodules. Within this region is a mirror repeat sequence that is potentially capable of forming an H-DNA structure. These results indicate that PEPC-7 has a central role in nitrogen-fixing nodules and that regulation of transcription is an important determinant of its activity.

In a search for genes that are only expressed in fruit bodies of the basidiomycete Agaricus bisporus, two cDNAs, hypA and hypB that encode hydrophobins have been isolated previously. In this study, the structure of hypB is resolved and it is shown that the two genes are differentially expressed, indicating that the encoded hydrophobins serve different functions in A. bisporus mushrooms. hypB encodes a polypeptide (HYPB) of 119 aa that shows little sequence identity with HYPA apart from the characteristic arrangement of eight cysteines found exclusively in hydrophobins. The temporal and spatial expression of the two hydrophobin-encoding genes during fruit body development was compared using Northern analysis and in situ hybridization. Accumulation of hypA mRNA was found in tissue fractions consisting of undifferentiated white hyphae. In situ hybridization showed that the highest hypA mRNA levels are not found in the outermost cell layers of the pileipellis but in the cell layers adjacent to that. The highest level of expression of hypB occurs early in development when the primordium differentiates into densely packed, randomly oriented cap hyphae and loosely packed, vertically oriented stipe hyphae. In mature mushrooms, a strong accumulation of hypB transcripts was found only in the transitional zone between cap and stipe tissue, demonstrating that transcription regulation of hype is clearly distinct from hypA. I I

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A rapid and inexpensive DNA diagnostic platform for fingerprinting Cdn. registered, wheat varieties has been developed. Two current, real-time applications being used in Canada include the determination of purity (% contamination) of grain shipments in rail cars and the monitoring of field plots that represent a midge varietal blend. The quantification of a sample is accomplished by fingerprinting single seeds and a sample having a mixture of varieties can be assayed. The Rapid ID Technology (RIDT) platform enables high-speed, high-throughput, robotic labautomation and low-cost single nucleotide polymorphism (SNP)-DNA fingerprinting in wheat. The inexpensive seed DNA extraction method, rapid PCR amplification, and miniaturization of the Invader assay for SNP scoring are all paramount for fingerprinting millions of single seeds per year in numerous rail cars/field plots at a low cost. The cost factor is the dominant concern in any fingerprinting program and the total cost for consumables for a single marker tested is estimated at less than $0.60 per seed. The RIDT platform can also segregate closely related cultivars, an important characteristic since many varieties have a common genetic background. The results ''from seed to fingerprint data'' at a central lab can then be transferred electronically to any location using a laboratory information management system. ( JALA 2009;14:221e231)