Wipaporn Wanthanalert and Mariena Ketudat-Cairns

pp. 23 - 32

Abstract

This research attempted to study the function of Os1bglu1 by RNAi technique. The suppression of Os1bglu1gene
was done using the 3’UTR region. The target gene fragment was cloned into the pHELLSGATE8 vector. The high percentages
of effective callus induction of 93% were obtained when the seeds were cultured on N6D medium for 4-6 weeks at 28°C. The
suitable transformation conditions were to incubate the calli with Agrobacterium (OD₆₀₀ = 0.02) and blot dry to remove excess
bacteria cells, then transferred to co-cultivation medium (pH 5.2) with 200 μM acetosyringone and incubate for three days at
25°C. The 20% transformation efficiency was obtained from the transformed calli with control plasmid, while transformation
efficiency of only 15% was obtained from pHELLSGATE8 Os1bglu1 constructs. The transformed calli with control construct
showed higher growth rate than the transformed calli with pHELLSGATE8 Os1bglu1construct. The expression of Os1bglu1
mRNA was not found in the transformed calli and siRNAs were found in the transformed calli. However no siRNAs were
detected in the control transformed calli. The regeneration efficiencies of 6% were obtained from only the calli transformed
with the control construct. The calli transformed with the knock down Os1bglu1 constructs were not able to regenerate. This
may indicated that Os1bglu1 is involved in regeneration of rice from callus tissue.