Generating Targeted Gene Knockout Lines in Physcomitrella patens to Study Evolution of Stress-Responsive Mechanisms.

The moss Physcomitrella patens possesses highly efficient homologous recombination allowing targeted gene manipulations and displays many features of the early land plants including high tolerance to abiotic stresses. It is therefore an invaluable model organism for studies of gene functions and comparative studies of evolution of stress responses in plants. Here, we describe a method for generating targeted gene knockout lines in P.

Regulation of plant gene expression by alternative splicing.

AS (alternative splicing) is a post-transcriptional process which regulates gene expression through increasing protein complexity and modulating mRNA transcript levels. Regulation of AS depends on interactions between trans-acting protein factors and cis-acting signals in the pre-mRNA (precursor mRNA) transcripts, termed 'combinatorial' control. Dynamic changes in AS patterns reflect changes in abundance, composition and activity of splicing factors in different cell types and in response to cellular or environmental cues.

Alternative splicing in plants.

The impact of AS (alternative splicing) is well-recognized in animal systems as a key regulator of gene expression and proteome complexity. In plants, AS is of growing importance as more genes are found to undergo AS, but relatively little is known about the factors regulating AS or the consequences of AS on mRNA levels and protein function. We have established an accurate and reproducible RT (reverse transcription)-PCR system to analyse AS in multiple genes.

Monitoring changes in alternative precursor messenger RNA splicing in multiple gene transcripts.

Alternative splicing (AS) increases the proteomic and functional capacity of genomes through the generation of alternative mRNA transcripts from the same gene. AS is now estimated to occur in a third of Arabidopsis and rice genes, and includes genes involved in the control of growth and development, responses to stress and signalling. Regulation of AS reflects the interactions between positive and negative cis sequences in the precursor messenger RNA and a range of trans-acting factors.