Gene Expression
Nebulas
Gene Expression NebulasSummary: Purpose: Comprehensive comparison of gene-expression profiles between 3mlpa (mutant with low phytic acid) and 3MWT (non-mutant with normal phyic acid) soybean lines (Glycine max) at five stages of seed development using RNA-Seq approaches. Methods: mRNA sequencing reads (101SE) were generated in triplicate from five seed developmental stages of 3mlpa and 3MWT soybean line using Illumina HiSeq 2000. Results: A total of 4235 differentially expressed genes, including 512-transcription factor genes were identified. Eighteen biological processes such as apoptosis, glucan metabolism, cellular transport, photosynthesis and 9 transcription factor families including WRKY, CAMTA3 and SNF2 were enriched during seed development. Genes associated with apoptosis, glucan metabolism, and cellular transport showed enhanced expression in early stages of lpa seed development, while those associated with photosynthesis showed decreased expression in late developmental stages. Conclusion: This study provides a global perspective of transcriptomal changes during soybean seed development in 3mlpa mutant. The results suggest that low phtic acid-causing mutations in 3mlpa play a role in inducing and suppressing plant defense responses during early and late stages of seed development, respectively.
Overall Design: RNA-Seq of five seed developing soybean seeds from mips1/mrp-l/mrp-n triple mutant line (with low phytic acid) and MIPS1/MRP-L/MRP-N non-mutant line (with normal phytic acid). The mRNA libraries from three biological replicates of each sample were sequenced as 101 SE using Illumina HiSeq 2000.
| Strategy: |
|
| Species: |
|
| Tissue: |
|
| Height/Length/Weight: |
|
| Isolation_source: |
|
| Development Stage: |
|
| Growth Protocol: | For each of the two experimental lines, 48 plants were grown in 12 pots (four plants per pot) containing Metro-Mix® 360 (Sun Gro) soilless media, over-layered with topsoil GardenPro ULTRALITE. All plants were grown in the same growth chamber unit, with controlled conditions, as follows: 14/10 h (day/night) photoperiod, 24/16 °C (day/night), light intensities in the range of 300 and 400 μE and 50–60 % relative humidity. About 41–47 plants from each experimental line were used for sampling developing seeds. |
| Treatment Protocol: | Seed length was the criterion for sampling different developmental stages. Pods were randomly selected, opened and, seed length was measured. Five developmental stages were defined by seed length as: (Stage 1) between 2-4 mm; (Stage 2) between 4-6 mm; (Stage 3) between 6-8 mm; (Stage 4) between 8-10 mm; and, (Stage 5) between 10-12 mm. Three biological replicates for each stage were taken, where each replicate sample was represented by a minimum of 10-15 seeds (stages 1-2), and at least 3 seeds (stages 3-5), collected from different pods on separate plants. Sampled seeds were immediately frozen in liquid nitrogen and stored at -70 °C. |
| Extract Protocol: | Whole soybean developing seeds sampled in triplicate from 3mlpa and 3MWT lines at five stages of seed development, were flash frozen, and total RNA was extracted using QIAGEN RNeasy Plant Mini Kit, with on column DNase digestion. |
| Library Construction Protocol: | High quality total RNA samples were used to generate 30 mRNA libraries using standard Illumina TruSeq RNA sample preparation kit protocol. |
| Molecule Type: | poly(A)+ RNA |
| Library Source: | |
| Library Layout: | SINGLE |
| Library Strand: | Forward |
| Platform: | ILLUMINA |
| Instrument Model: | Illumina HiSeq 2000 |
| Strand-Specific: | Specific |
| Data Resource | GEN Sample ID | GEN Dataset ID | Project ID | BioProject ID | Sample ID | Sample Name | BioSample ID | Sample Accession | Experiment Accession | Release Date | Submission Date | Update Date | Species | Race | Ethnicity | Age | Age Unit | Gender | Source Name | Tissue | Cell Type | Cell Subtype | Cell Line | Disease | Disease State | Development Stage | Mutation | Phenotype | Case Detail | Control Detail | Growth Protocol | Treatment Protocol | Extract Protocol | Library Construction Protocol | Molecule Type | Library Layout | Strand-Specific | Library Strand | Spike-In | Strategy | Platform | Instrument Model | Cell Number | Reads Number | Gbases | AvgSpotLen1 | AvgSpotLen2 | Uniq Mapping Rate | Multiple Mapping Rate | Coverage Rate |
|---|