Computational protocol: S phase of the cell cycle: a key phase for the regulation of thermodormancy in barley grain

Similar protocols

Protocol publication

[…] In order to analyse cell-cycle transcription activity, the expression of related genes as markers of cell-cycle phase was studied. The TIGR barley expressed sequence tag (EST) databank (http://www.ncbi.nlm.nih.gov/blast/Blast.cgi?PAGE_TYPE=BlastSearch&PROG_DEF=blastn&BLAST_PROG_DEF=megaBlast&BLAST_SPEC=Plants_EST) was screened using specific protein motifs for each class to identify barley genes. To complete and confirm the databank annotation, specific protein motifs were identified using the translation software from infobiogene (http://www.ncbi.nlm.nih.gov/Structure/cdd/wrpsb.cgi). A class could be attributed for each gene with specific motifs, i.e. for CDKDA (CDKA1), CDKB (CDKB1), CDKD (CDKD1), CYCA (CYCA3), CYCB (CYCB1), and CYCD (CYCD4) (Vandepoele et al., 2002), and KRP (KRP4) and WEE (WEE1) (Sorrell et al., 2002). Specific primers were then designed using the software Primer3 for each EST (Supplementary Table S1) to perform reverse-transcription (RT) PCR. As an internal standard, a fragment of the barley actin gene (GI24496451) was used. Each expression profile presented was repeated at least three times in three independent experiments. […]

Pipeline specifications

Software tools BLASTN, Primer3
Application qPCR
Organisms Hordeum vulgare
Chemicals Abscisic Acid, Hydroxyurea