Molecular cloning and genetic mapping of perennial ryegrass casein protein kinase 2 α-subunit genes
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The α-subunit of the casein protein kinase CK2 has been implicated in both light-regulated and circadian rhythm-controlled plant gene expression, including control of the flowering time. Two putative CK2α genes of perennial ryegrass (Lolium perenne L.) have been obtained from a cDNA library constructed with mRNA isolated from cold-acclimated crown tissue. The genomic organisation of the two genes was determined by Southern hybridisation analysis. Primer designs to the Lpck2a-1 and Lpck2a-2 cDNA sequences permitted the amplification of genomic products containing large intron sequences. Amplicon sequence analysis detected single nucleotide polymorphisms (SNPs) within the p150/112 reference mapping population. Validated SNPs, within diagnostic restriction enzyme sites, were used to design cleaved amplified polymorphic sequence (CAPS) assays. The Lpck2a-1 CAPS marker was assigned to perennial ryegrass linkage group (LG) 4 and the Lpck2a-2 CAPS marker was assigned to LG2. The location of the Lpck2a-1 gene locus supports the previous conclusion of conserved synteny between perennial ryegrass LG4, the Triticeae homoeologous group 5L chromosomes and the corresponding segment of rice chromosome 3. Allelic variation at the Lpck2a-1 and Lpck2a-2 gene loci was correlated with phenotypic variation for heading date and winter survival, respectively. SNP polymorphism may be used for the further study of the role of CK2α genes in the initiation of reproductive development and winter hardiness in grasses.
KeywordsPerennial ryegrass Casein kinase 2 α-subunit Single nucleotide polymorphism Cleaved amplified polymorphic sequence Heading date Conserved synteny
This work was supported, in part, by Grants-in-Aid for Scientific Research (Nos. 14360160 and 17380163 to T.Y.) from the Ministry of Education, Science, Sports and Culture, Japan.The research in Australia was supported by Primary Industries Research Victoria and the Molecular Plant Breeding Cooperative Research Centre. The authors thank Prof. Michael Hayward (Rhydgoch Genetics, Aberystwyth, UK) and Drs. Aidyn Mouradov and Ulrik John for careful critical reading of the manuscript. The scientific advice and support of Prof. G. Spangenberg is gratefully acknowledged.
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