|
| Status |
Public on Mar 07, 2023 |
| Title |
Z4-2 |
| Sample type |
RNA |
| |
|
| Source name |
Portunus trituberculatus larvae
|
| Organism |
Portunus trituberculatus |
| Characteristics |
stages: 4
|
| Treatment protocol |
Samples were collected when 70%-80% of the population has molted to the desired stage. The samples were immediately rinsed with sterilized seawater, snap-frozen in liquid nitrogen, and then stored at -80 ℃. Three biological replicates of six stages were used for the following analysis.
|
| Growth protocol |
In short, rotifers (Branchionus plicatilis) 30-40 ind /mL and microalga (Platymonas subcordiformis) (5-10) ×104/mL were fed every two hours during Z1-2) while Artemia nauplii 2-3 ind /mL was provided to Z3-C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted by mirVanaTM RNA Isolation Kit (Applied Biosystem p/n AM1556 ) following the manufacturer's instructions.
|
| 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.
|
| |
|
| Hybridization protocol |
0.6 μg of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/μg 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 Agilent Portunus trituberculatus Gene Expression（8*60K）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.
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505C) using one color scan setting for 4x180k array slides (Scan Area 61x21.6 mm, Scan resolution 3um, Dye channel is set to Green and Green PMT is set to 100%).
|
| Description |
zoea
|
| 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 13.0(Agilent). Probe that at least 1 out of 2 samples flagged as Detected were maintained.
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| |
|
| Submission date |
Mar 03, 2023 |
| Last update date |
Mar 08, 2023 |
| Contact name |
Chen Jiameng |
| E-mail(s) |
2011091076@nbu.edu.cn
|
| Organization name |
Ningbo University
|
| Street address |
No.169 Qixing South Road, Meishan Street, Beilun District, Ningbo City, Zhejiang Province, China
|
| City |
Ningbo |
| ZIP/Postal code |
315800 |
| Country |
China |
| |
|
| Platform ID |
GPL33207 |
| Series (1) |
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