Q-PCR LD (12:12) 17C ELF3

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BioDare ID 12962294335805
Author Aurora Pinas-Fernandez
Institution University of Edinburgh
License

Description

Seedlings were entrained for 7 days at 22C in 12:12 white light. Then shifted to 17C 12:12 white light. Samples were collected on the 6th day at 17 degrees.

Purpose

Experiment carry out to reparameterize the clock off Succrose

Comments

Sample time 24h. 24h sample lost for some genotypes Primer JF260/261 used for this experiment. Primers used in Millar lab and other Robust experiments No data available for Col, prr7-3, prr9-1 or prr7-3/prr9-1

Sample preparation

Samples were harvested and keeped on RNAlader for some weeks, at 4C and -20C. RNa extraction was done following protocol of GE Health Care-illustra RNAspin 96RNA Isolation kit. Samples were spect with Nanodrop. cDNA synthesis was done with Super Script Vilo kit( Invitrogene) from 1ugr RNA.

Measurement

Protocol

RT-PCR (Robot + Light Cycler 480)

JF260-ELF3-F: GGAAAGCCATTGCCAATCAA JF261-ELF3-R: ATCCGGTGATGCAGCAATAAGT JF588-ELF3-F:GGTCATTATATGCCTACACCA JF589-ELF3-F: GAAGTAGCCATTACCAGGAG

JF260-ELF3-F & JF261-ELF3-R JF588-ELF3-F & JF589-ELF3-F Cycle: 2 - Roche Parameters-cDNA dil 1/10

Experimental conditions

Entrainment 22C Diurnal

Growth on Agar 1.2% MS 0.5 for 7.0 days (Entrainment 22C Diurnal).

Light

Type Duration (days) Cycle (h) Start Duration Spectrum Source Intensity
diurnal light 7 24 0:00 12:00 white tube 60

Temperature

Type Duration (days) Cycle (h) Base (°C) Warm (°C) Warm start Warm duration
constant temperature 7 24 22 -- -- --

17C LD (12:12) Off Sucrose

Growth on Agar 1.2% MS 0.5 for 6.0 days (17C LD (12:12) Off Sucrose).

Light

Type Duration (days) Cycle (h) Start Duration Spectrum Source Intensity
diurnal light 6 24 0:00 12:00 white tube 60

Temperature

Type Duration (days) Cycle (h) Base (°C) Warm (°C) Warm start Warm duration
constant temperature 6 24 17 -- -- --

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