Genetic Control of Parthenogenesis in Kentucky Bluegrass: Results from a Sexual x Apomictic Cross

Apomixis, as it exists in Poa pratensis L., permits to combine genotype fixation with propagation by seed. The process of apomictic seed production involves the formation of embryo sacs without meiotic reduction (apospory) and embryos without egg cell fertilization (parthenogenesis). Further information on the genetic control of apomixis was obtained by analyzing aposporous parthenogenesis and the segregation of molecular markers in a progeny resulting from a “sexual” (S) x “apomictic”(A) cross. Data from the 35 F1 plants examined so far have shown that parthenogenesis segregated 1:1, confirming control by a dominant gene, simplex in the parthenogenetic parent. Analysis of variance and regression were used to find single markers from the A and S parents affecting the trait in the 15 parthenogenetic progenies. A minimum of 4 genes from the sexual parent and 1 from the apomictic parent influencing the expression of parthenogenesis appeared to segregate in this cross

Identification and validation of reference genes for quantitative RT-PCR normalization in wheat

<p>Abstract</p> <p>Background</p> <p>Usually the reference genes used in gene expression analysis have been chosen for their known or suspected housekeeping roles, however the variation observed in most of them hinders their effective use. The assessed lack of validated reference genes emphasizes the importance of a systematic study for their identification. For selecting candidate reference genes we have developed a simple <it>in silico </it>method based on the data publicly available in the wheat databases Unigene and TIGR.</p> <p>Results</p> <p>The expression stability of 32 genes was assessed by qRT-PCR using a set of cDNAs from 24 different plant samples, which included different tissues, developmental stages and temperature stresses. The selected sequences included 12 well-known HKGs representing different functional classes and 20 genes novel with reference to the normalization issue. The expression stability of the 32 candidate genes was tested by the computer programs geNorm and NormFinder using five different data-sets. Some discrepancies were detected in the ranking of the candidate reference genes, but there was substantial agreement between the groups of genes with the most and least stable expression. Three new identified reference genes appear more effective than the well-known and frequently used HKGs to normalize gene expression in wheat. Finally, the expression study of a gene encoding a PDI-like protein showed that its correct evaluation relies on the adoption of suitable normalization genes and can be negatively affected by the use of traditional HKGs with unstable expression, such as actin and α-tubulin.</p> <p>Conclusion</p> <p>The present research represents the first wide screening aimed to the identification of reference genes and of the corresponding primer pairs specifically designed for gene expression studies in wheat, in particular for qRT-PCR analyses. Several of the new identified reference genes outperformed the traditional HKGs in terms of expression stability under all the tested conditions. The new reference genes will enable more accurate normalization and quantification of gene expression in wheat and will be helpful for designing primer pairs targeting orthologous genes in other plant species.</p

The PDI (Protein Disulfide Isomerase) gene family in wheat.

The PDI (Protein Disulfide Isomerase) gene family includes several members whose products are responsible for diversified metabolic functions. PDI and PDI-like proteins differ for number and position of thioredoxin-like (TRX-like) active (a type) and inactive (b type) domains, for presence/absence of other domains and of the KDEL signal of retention in the endoplasmic reticulum (ER). The phylogenetic analysis of typical PDI and PDI-like protein sequences resolved them into 10 groups (1), 5 of them (I-V) had 2 TRX-like active domains, whereas the remaining ones owned only a single TRX-like active domain (VI-VIII, QSOX and APRL). In particular, QRX and APRL were not included in this study due to their putative non-isomerase enzymatic activities encoded by an additional domain. The aim of the present research was the study of the complexity and diversity of the PDI gene family in wheat, with particular focus on the genes encoding PDIlike proteins structurally similar to TaPDIL1-1 (group I), the first identified and best characterized member of the PDI family, also named typical PDI. The most important function of typical PDI is the formation and isomerization of disulfide bonds during protein folding, which are accomplished by its two active TRX-like sites sharing the characteristic tetrapeptide –CGHC-. Several studies of molecular characterization, expression analysis and cell localisation in rice and maize have suggested the involvement of typical PDI in the assembly and deposition of storage proteins in these species (2, 3, 4). The characterization and chromosome location of the three homoeologous gene sequences encoding typical PDI and of their promoter sequences have been reported previously (5)

Relationship between the D genome of hexaploid wheats (AABBDD) and Ae. squarrosa as deduced by seed storage proteins and molecular marker analyses

The electrophoretical analyses of seed storage protein components from the gliadin and glutenin fractions in T. aeslivum ssp. vulgare, compaction, sphaerococcum, macha, vavilovii, and spelta have revealed limited variation at the tightly linked coding loci Gli-D1/Glu-D3, and Glu-D1, located respectively on the short and long arm of chromosome ID, and at the GH-D2 locus, positioned on the short arm of chromosome 6D. Much higher variation was observed, for the same protein components, in the wild diploid Ae. squarrosa, the D genome donor of the aestivum group. Genetic variation in the same wheat subspecies and in Ae. squarrosa has also been evaluated by Southern hybridization of genomic DNAs, which were digested with several restriction enzymes, and hybridized with cloned sequences of genes coding for seed storage proteins. The much higher degree of variation observed for the seed storage protein genes of Ae. squarrosa, in comparison with the variation exhibited by the proteins encoded by the D genome chromosomes of hexaploid wheats, supports the hypothesis that a limited number of crosses gave rise to hexaploid wheats of the aestivum group

The Protein Disulfide Isomerase gene family in bread wheat (T. aestivum L.)

<p>Abstract</p> <p>Background</p> <p>The Protein Disulfide Isomerase (PDI) gene family encodes several PDI and PDI-like proteins containing thioredoxin domains and controlling diversified metabolic functions, including disulfide bond formation and isomerisation during protein folding. Genomic, cDNA and promoter sequences of the three homoeologous wheat genes encoding the "typical" PDI had been cloned and characterized in a previous work. The purpose of present research was the cloning and characterization of the complete set of genes encoding PDI and PDI like proteins in bread wheat (<it>Triticum aestivum </it>cv Chinese Spring) and the comparison of their sequence, structure and expression with homologous genes from other plant species.</p> <p>Results</p> <p>Eight new non-homoeologous wheat genes were cloned and characterized. The nine PDI and PDI-like sequences of wheat were located in chromosome regions syntenic to those in rice and assigned to eight plant phylogenetic groups. The nine wheat genes differed in their sequences, genomic organization as well as in the domain composition and architecture of their deduced proteins; conversely each of them showed high structural conservation with genes from other plant species in the same phylogenetic group. The extensive quantitative RT-PCR analysis of the nine genes in a set of 23 wheat samples, including tissues and developmental stages, showed their constitutive, even though highly variable expression.</p> <p>Conclusions</p> <p>The nine wheat genes showed high diversity, while the members of each phylogenetic group were highly conserved even between taxonomically distant plant species like the moss <it>Physcomitrella patens</it>. Although constitutively expressed the nine wheat genes were characterized by different expression profiles reflecting their different genomic organization, protein domain architecture and probably promoter sequences; the high conservation among species indicated the ancient origin and diversification of the still evolving gene family. The comprehensive structural and expression characterization of the complete set of <it>PDI </it>and <it>PDI</it>-like wheat genes represents a basis for the functional characterization of this gene family in the hexaploid context of bread wheat.</p