Differential LongSAGE tag abundance analysis in a barley seed germination time course and validation with relative real-time RT-PCR

White, Jessica, Pacey-Miller, Toni, Bundock, Peter and Henry, Robert (2008) Differential LongSAGE tag abundance analysis in a barley seed germination time course and validation with relative real-time RT-PCR. Plant Science, 175 6: 858-867.


Author White, Jessica
Pacey-Miller, Toni
Bundock, Peter
Henry, Robert
Title Differential LongSAGE tag abundance analysis in a barley seed germination time course and validation with relative real-time RT-PCR
Journal name Plant Science   Check publisher's open access policy
ISSN 0168-9452
Publication date 2008-12
Sub-type Article (original research)
DOI 10.1016/j.plantsci.2008.08.008
Volume 175
Issue 6
Start page 858
End page 867
Total pages 10
Place of publication Ireland
Publisher Elsevier Ireland Ltd
Language eng
Formatted abstract A Long Serial Analysis of Gene Expression (LongSAGE) approach was employed to identify changes in mRNA transcript abundance in a time course of malting barley (Hordeum vulgare L.). Statistical analyses confidently identified 57 LongSAGE sequence tags as having significant changes in abundance between points in the time course. Eight of the genes which correspond to these tags were targeted for validation by relative real-time reverse transcriptase PCR (RT-PCR) analysis. Each gene was analysed by SYBR® Green detection in grain sampled at each of four time points from dry seed to grain germinated to 120 h post-steeping. Among the genes examined are alpha-amylase type B, a key starch degrading enzyme involved in germination (1-3,1-4)-beta-d-glucanase, the major cell wall degrading enzyme, and cysteine proteinase EP-B, an important enzyme of proteolysis in barley seed germination. These three transcripts show significant up-regulation at 48 h post-steeping and remain significantly elevated throughout malting. These results provide transcriptional data to support the understanding of how the relative rates of protein and carbohydrate modification contribute to malting and brewing. mRNA abundance levels observed using real-time RT-PCR show good correlation with the data obtained from the LongSAGE study. This confirms the sensitivity of detection obtainable with SAGE technology and validates the patterns of change of transcript abundance exhibited by some key genes for barley seed germination.
Keyword Germination
Hordeum Vulgare
Real-time Reverse Transcriptase Pcr
Long Serial Analysis Of Gene Expression (longsage)
Q-Index Code C1
Q-Index Status Provisional Code
Institutional Status Unknown

Document type: Journal Article
Sub-type: Article (original research)
Collections: Queensland Alliance for Agriculture and Food Innovation
ERA 2012 Admin Only
 
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