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

methods

https://eprints.lib.hokudai.ac.jp/dspace/bitstream/2115/864/1/TAG112-1.pdf

Spangenberg, G. (Ed.): Molecular Breeding of Forage Crops

Molecular cloning and genetic mapping of perennial ryegrass casein protein kinase 2 α-subunit genes

To identify potential candidates for acquiring stress tolerance, a new annealing control primer (ACP) system was used to identify the differentially expressed genes. Alfalfa (Medicago sativa L.) seedlings were exposed to various abiotic stresses such as cold (4°C for 6 h), heat (42°C for 6 h), salt (300 mM for 6 h), drought (withdrawing irrigation for 48 h), copper (500 μM for 6 h), cadmium (500 μM for 6 h), and arsenic (500 μM for 6 h). Primer sets 41 and 93 were differentially expressed and identified as same sequence, which represents a mitochondrial small heat-shock protein encoding gene, MsHsp23. This band was markedly increased or induced in alfalfa under heat, salt, and arsenic stresses. Differential expression of MsHsp23 was further evaluated by Northern blot analysis. Temporal expression analysis showed that mRNA pool was altered as early as 1 h of treatment. Thus, differential accumulation of MsHsp23 under heat, salt, and arsenic stresses suggests its potential involvement in diverse abiotic stress tolerance, and thereby making a target for further molecular analysis.

background

/pdf/identification-of-mshsp23-gene-using-annealing-control-4d0ma8pc0y.pdf

Spangenberg, G. (Ed.): Molecular Breeding of Forage Crops

Citations

Molecular cloning and genetic mapping of perennial ryegrass casein protein kinase 2 α-subunit genes

Identification of MsHsp23 gene using annealing control primer system

References

Molecular cloning and genetic mapping of perennial ryegrass casein protein kinase 2 α-subunit genes

Identification of MsHsp23 gene using annealing control primer system

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Molecular breeding of forage crops : proceedings of the 2nd Inernational Symposium, Molecular Breeding of Forage Crops, Lorne and Hamilton, Victoria, Australia, November 19-24, 2000

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