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Status |
Public on Jan 01, 2015 |
Title |
Leaf-ZH2_spring_rep2 |
Sample type |
RNA |
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Source name |
fresh shoots, Zhonghuang 2, replicate 2
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Organism |
Camellia sinensis |
Characteristics |
tissue: fresh shoots (two and a bud) sampling season: spring
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Treatment protocol |
In the spring, the fresh shoots (two and a bud) were sampled from the four cultivars.
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Growth protocol |
Four tea plant cultivars, "Longjing 43", "Zhongcha 108", "Zhonghuang 1" and "Zhonghuang 2" were grown in the China National Germplasm Hangzhou Tea Repository (CNGHTR) at the Tea Research Institute, Chinese Academy of Agricultural Sciences (TRI, CAAS).
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Extracted molecule |
total RNA |
Extraction protocol |
RNA was extracted by Trizol (Life Technology, 15596-018) following the manufacturer's.
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Label |
Cy3
|
Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 0.2 μg RNA using the One-Color Low RNA Input Linear Amplification PLUS kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
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Hybridization protocol |
0.6 μg of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 22.5μl containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers' instructions. On completion of the fragmentation reaction, 22.5μl of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to the Camellia sinensis 44K custom array (Agilent) for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent), then dried immediately by brief centrifugation.
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Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner using one color scan setting.
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Description |
Gene expression in spring
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Data processing |
The scanned images were analyzed with Feature Extraction Software 10.7.1.1 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities as the raw data. Raw data were normalized in quantile algorithm with Genespring 12.0(Agilent). Probe that at least 1 out of 2 samples flagged as "Detected" were maintained.
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Submission date |
Nov 11, 2013 |
Last update date |
Jan 01, 2015 |
Contact name |
Lu Wang |
E-mail(s) |
wanglu317@tricaas.com
|
Organization name |
Tea Research Institute, Chinese Academy of Agricultural Sciences
|
Street address |
9th, South Meiling Road
|
City |
Hangzhou |
State/province |
Zhejiang |
ZIP/Postal code |
310008 |
Country |
China |
|
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Platform ID |
GPL17911 |
Series (1) |
GSE52255 |
Development of a 44K custom oligo microarray using 454 pyrosequencing data for large scale gene expression analysis in tea plant (Camellia sinensis) |
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