Rapid assessment of single-copy nuclear DNA variation in diverse species

We investigated the use of PCR primers designed to conserved exons within nuclear DNA to amplify potentially variable regions such as introns or hypervariable exons from a wide range of species. We then explored various approaches to assay population-level variation in these PCR products. Primers designed to amplify regions within the histone H2AF, myoglobin , MHC DQA , and aldolase (ALD) genes gave clean amplifications in diverse mammals (DQA) , and in birds, reptiles and mammals ( aldolase, H2AF, myoglobin ). The sequenced PCR products generally, but not always, confirmed that the correct locus had been amplified. Several primer sets produced smaller size fragments consistent with preferential amplification of intronless pseudogenes; this was confirmed by sequencing seal and reptile H2AF PCR products. Digestion with randomly selected four-base recognizing enzymes detected variation in some cases but not in others. In species/gene combinations with either low (e.g. seal H2AF, ALD-A ) or high (e.g. skink ALD-1 ) nucleotide diversity it was more efficient to sequence a small number of distantly related individuals (e.g. one per geographic population) and from these data to identify informative or potentially informative restriction enzymes for 'targeted' digestion. We conclude that for studies of population-level variation, the optimal approach is to use a battery of primers for initial PCR of both mtDNA and scnDNA loci, select those that give clean amplifications, and sequence one sample from each population to (i) confirm gene identity, (ii) estimate the amount of variation and, (iii) search for diagnostic restriction sites. This will allow determination of the most efficient approach for a large-scale study.     

Growth Responses of Dwarf Bean to Infection by Pseudomonas phaseolicola (Burk) Dows

At an early growth stage inoculation of both unifoliates ofdwarf beans with Pseudomonas phaseolicola (Burk) Dows. Race1 inhibited trifoliate production, but inoculation of one unifoliateonly reduced subsequent leaf expansion, and the opposite uninfectedunifoliate became significantly larger than unifoliates on uninfectedplants. Inoculation of the first or second trifoliates or thecotyledonary node at a later developmental stage all reducedprimary-shoot growth by 25 to 30 per cent, but there were markeddifferences in the extent of secondary-shoot development inleaf axils below the inoculation point. The growth effects were related to the distribution and activityof the bacterial toxin within the plant, particularly to differencesin the rate of invasion of apical regions. The primary toxineffects appear to be on apical function and leaf expansion.     

Micronuclear and Macronuclear Sequences of a Euplotes crassus Gene Encoding a Putative Nuclear Protein Kinase

The sequences of a 1.8-kbp macronuclear DNA molecule (V3), and the majority of its micronuclear counterpart, are reported. The macronuclear V3 DNA molecule contains an open reading frame that is interrupted by a single intron, while the micronuclear copy is interrupted by four internal eliminated sequences, one of which is located within the intron. The predicted protein product of the macronuclear V3 gene is a 471-amino acid polypeptide that is very similar to a group of protein-serine/threonine kinases from both plant and animal species, some of whose members appear to be involved in cell cycle or growth control.     

Polymorphic microsatellite loci in peafowl (Pavo cristatus)

Understanding how the genetic characteristics of parents influence reproductive output is central to predicting the dynamics of small, endangered populations. We conducted a breeding experiment to look at the paternal genetic effects on offspring sex, fertility and growth in the peafowl (Pavo cristatus). Microsatellite loci were developed to allow maternity assignment and thus to allow us to separate maternal from paternal effects. We found 19 polymorphic loci in our inbred, captive population, six of which were only slightly polymorphic (H_E range: 0.04–0.70). The remaining 13 loci were polymorphic enough to determine maternity by exclusion in approximately 85% of offspring.