Comparative Transcriptome Analysis of Abiotic Stress Tolerance

Given that SR3 has demonstrated improved levels of tolerance to both salinity and drought, we proposed to compare the sequence variation of ESTs that are produced under the control and after exposure to saline and osmotic stresses to gain the association between genetic variation and abiotic stress tolerance. Three-leaf-stage seedlings of both cv. SR3 and cv. JN177 were exposed to half strength Hoagland’s liquid medium containing either 200 mM NaCl for 0, 0.5, 1, 12, 24, 48 or 72 h, or to 18 % PEG for 0, 0.5, 1, 12, 24 or 48 h. RNA was extracted from both roots and leaves using the Trizol reagent (Invitrogen, USA), and purified using an OligotexTM-dT₃₀ mRNA Purification Kit (TaKaRa, Japan). Equimolar amounts of the various RNA samples were mixed to obtain both an SR3 and a JN177 mRNA pool, which formed the template for the construction of two cDNA libraries using a CloneMiner™ cDNA Library Construction Kit (Invitrogen, USA), according to the manufacturer’s protocol. Given that the genetic variation induced by asymmetric somatic hybridization was remarkably higher than that induced by other mutagenic factors protoplast formation, UV radiation, callus induction and plant regeneration [31 (link), 32 (link)], other cDNA libraries reflecting the genetic variation of these mutagenic factors were not constructed.