| dc.description.abstract | Abstract
There are 9 members of the class I small heat shock proteins (sHSP-CI) gene family in rice (Oryza sativa L. cv. Tainong No.67), of which on chromosome 1 and chromosome 3. Interestingly, Oshsp17.3 and Oshsp18.0 on chromosome 3 are linked head-to-head and share a 356-bp putative bi-directional promoter. A possible azetidine (Aze)-responsive element (AZRE;GTCCTGGACG) on Oshsp17.3 promoter was found to be capable of directing expression in response to Aze treatment. The truncation of AZRE decreased the induction of both Aze induction and heat shock (HS) induction. To improve high control value, we constructed a transient expression system using Arabidopsis protoplast to assay Oshsp17.3 promoter. The results showed that there was not significant difference for HS induction and Cd induction. So we use stable transformation to confirm the role of AZRE and AT-rich region. Five constructs ( p567(－): full length of Oshsp17.3 promoter; p567(－)ΔAZRE: AZRE truncation of Oshsp17.3 promoter; p567(＋): full length of Oshsp18.0 promoter; P1600: full length of Oshsp16.9A promoter; P500: AT-rich region truncation of Oshsp16.9A promoter) were generated and introduced into Arabidopsis plants via Agrobacterium. Totally 33 independent lines were constructed for further study. The copy number of transgene was estimated by southern-blot hybridization. One or two copies of transgenic independent line was selected and characterized under stress.
Different stage of Arabidopsis transgenic plants were exposed to HS and Aze treatment for histochemical analysis. Two-day-old seedlings and of control and transgenic plants showed constitutive blue GUS staining under stress. Analysis of 14-day-old seedlings revealed that p567(＋), p567(－), and p567(－)ΔAZRE of T2 transgenic plants showed blue GUS staining under HS and Aze stress, but no or less
detectable blue GUS staining was found under normal growth conditions. On the other hand, P1600 and P500 transgenic plants showed dominant blue GUS staining under HS and Aze stress. For analysis of one-month-old seedlings of T2 transgenic plants, all tissues in p567(＋) and p567(－) have blue GUS staining under HS and Aze treatment. In contrast, blue GUS staining was detected in HS treated p567(－)ΔAZRE but not in Aze treated p567(－)ΔAZRE.
Analysis of GUS activity of the transgenic plants in response to stress induction indicated that GUS activity of p567(＋) and p567(－) increased under HS and Aze treatment. In contrast, p567(－)ΔAZRE showed low expression level of GUS activity under HS stress, and GUS activity was not detected in Aze treated p567(－)ΔAZRE transgenic plants. On the other hands, as we would expect that P1600 showed GUS activity in response to HS but no GUS activity under Aze treatment. Besides, no GUS activity was detected in P500 under HS and Aze stress. Therefore we confirm the role of AZRE on the sHSP-CIs promoter using in vivo system.
To improve stress tolerance of Phalaenopsis sp. to environmental stresses, we also try to transform stress genes into this important flower plants in Taiwan. A rice sHSP-CI gene (Oshsp16.9A) and a barley (Hordeum vulgare L.) group 3 LEA protein gene (HVA1) were introduced into Phalaenopsis via Agrobacterium. The works for gene transformation into Phalaenopsis were accomplished. Analysis of stress tolerance conferred by Oshsp16.9A and HVA1 in Phalaenopsis needs further study to characterize in the future. | en_US |