Biology Reference
In-Depth Information
(Invitrogen, Carlsbad, US) and dT
18
VN anchor primers. LightCycler 480 SYBR
Green I Master kit (Roche Diagnostics, Indianapolis, US) was used to perform 15
µ
l real-time RT-PCR reactions in 384-well plates according to manufacturer's in-
structions by using Light Cycler 480 real-time PCR system (Roche Diagnostics).
PCR primers with T
m
value of 60°C were used (Table 5). Three biological repli-
cates were analysed for shoot apex samples from different developmental stages
(Figure 4), and two biological replicates were used for pooled shoot apex and leaf
samples (Table 6).
Authors' Contributions
TH, KM and PE designed all experiments. PE coordinated the study and helped
to draft the manuscript. TH run the real-time PCR analysis, performed flower-
ing gene searches from sequence databases, and drafted the manuscript together
with KM. KM constructed the subtracted cDNA libraries and performed bioin-
formatics analysis together with KF. KF also helped to draft the manuscript. MR
participated in flowering time analysis and sampling of shoot apices. PA and LP
were responsible for the EST sequencing. All authors read and approved the final
manuscript.
Acknowledgements
Dr. Michael Brosche is acknowledged for his kind help in the production of sub-
tracted cDNA libraries and M.Sc. Techn. Erkko Airo for his valuable technical
help. In addition, Finnish Ministry of Agriculture and Forestry is thanked for
financial support.
References
1. Ausín I, Alonso-Blanco C, Martinez-Zapater M: Environmental regulation of
flowering. Int J Dev Biol 2005, 49:689-705.
2. Putterill J, Laurie R, Macknight R: It's time to flower: the genetic control of
flowering time. Bioessays 2004, 26:363-373.
3. Simpson GG: The autonomous pathway: epigenetic and post-transcriptional
gene regulation in the control of Arabidopsis flowering time. Curr Opinion
Plant Biol 2004, 7:570-574.
4. Imaizumi T, Kay SA: Photoperiodic control of flowering: not only by coinci-
dence. Trends Plant Sci 2006, 11:550-558.
