Transcriptome sequencing and microarray design for functional genomics in the extremophile Arabidopsis relative Thellungiella salsuginea

ABSTRACT: Arabidopsis thaliana is not particularly stress tolerant and may lack protective mechanisms required to survive extreme environmental conditions. Thellungiella salsuginea has therefore attracted increasing interest as an alternative plant model species that possesses high tolerance of various abiotic stresses. While the T. salsuginea genome has recently been sequenced, its annotation is still far from complete and, due to the low number of ESTs available, evidence for actual transcriptional activity is lacking for most genes. ESTs were generated from a non-normalized and a normalized library synthesized from RNA pooled from plant samples from different developmental stages and grown under abiotic stress conditions. The ESTs of Thellungiella was sequenced by 454 pyrosequencing method. More than 1 million sequence reads were assembled into 42,810 unigenes, approximately 50% of which could be functionally annotated. From this sequence information, we constructed a 44k Agilent oligonucleotide microarray. A comparison of same-species and cross-species hybridization results showed the superior performance of the newly designed array for Thellungiella samples. In addition, the array was validated in a cold acclimation experiment that used the appropriately adapted MapMan software for analysis and visualization. To examine the quality of the Thellungiella array and to assess whether hybridization results from a dedicated array are superior to cross-species hybridization using the Arabidopsis array from the same manufacturer, we performed reciprocal array hybridizations. In this experiment, RNA from the Yukon accession of Thellungiella and RNA from the Col-0 accession of Arabidopsis were both hybridized on the Agilent Thellungiella and Arabidopsis expression arrays. For both species, RNA was extracted from acclimated and non-acclimated plants from three independent biological replicates.

ORGANISM(S): Eutrema salsugineum

SUBMITTER: Dirk Hincha 

PROVIDER: E-GEOD-42156 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress