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To screen for putative apomictic embryos possibly due to introduction of a key gene into a fully functional sexual background the Single Sequence Length Polymorphism (SSLP) assay was used to detect plants heterozygous for up to 15 markers. F2 populations of AtDMC1: AtSERK1 overexpressing plants generated during year 1 and 2 were screened for putative apomictic seeds using the SSLP analysis. The population showed segregation pattern similar to the wt. The highest number of heterozygous SSLP markers in a single plant found in this population was 15 and the chance to get this plant by chance only was more then 1/4000. Thus the procedure is sensitive enough to find as low is 0.1 % of potential apomicts within an otherwise sexually propagated population of plants.
A major objective of WP6 is to deliver novel germplasm of Poa pratensis expressing newly identified candidate genes derived in WP1. Using methodology developed in Year 1, a number of different P.pratensis germplasm provided by partner 3 were screened for responsiveness in tissue culture. Stable transformants of P.pratensis have been generated, using germplasm A24 and test constructs expressing uidA, and the presence of the introduced transgenes confirmed in T0 transformants. To complement transformation of apomictic P.pratensis, a number of different sexual P. pratensis were tested for responsiveness to tissue culture; in all germplasm screened responsivenes of sexual P.pratensis was low and stable transformation could not be detected.
The most consistently responsive sexual germplasm identified were derivatives of the S3 line. Transition to reproductive growth appears to be strongly inhibited in P.pratensis that has undergone regeneration through tissue culture. Plants will continue to be examined for flowering after vernalisation treatments in subsequent years as part of the continued analysis of this material after the end of ApoTool.
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