| Drought is one of the most widespread abiotic constraints to maize production in developing countries.Significant difference of drought tolerance has been found among maize inbred lines.However,Drought tolerance is believed to be the result of cooperative interactions among multiple morphological,physiological and biochemical characters. Moreover,different genotypes may have different responses to drought stress.Along with interference of environment,it is usually difficult to evaluate drought tolerance under ordinary conditions.The identification of candidate genes is helpful to improve the efficiency of breeding for increased drought tolerance by marker-assisted selection and to improve drought tolerance of maize varieties.In previous study,we identified 3 tolerant(81565,N87-1 and R09) and 2 sensitive (200B and ES40) inbred line from 57 parent lines of commercial maize hybrids under strict drought stress.Based on the results,the method of mRNA differential display was used to isolate differential expression genes from a drought tolerant maize inbred line '81565' under drought stress and well-watered control.Two down-regulated expression gene fragments(MD1 and MD2) and one up-regulated expression gene fragment were obtained under drought stress.Sequence analysis and homology alignment showed that MD2 had 88%similarity with gene PP2C,encoding serine / threonine phosphorylase 2C in extremely drought tolerant Sporobolus stapfianus.The full length cDNA sequence of gene fragment MD2 was cloned with the method of in silicon cloning and reverse transcription PCR(RT-PCR).The result indicated that the full length cDNA sequence cloned is a new member of PP2C gene family in maize,and its open reading frame of 1164 bp encodes 388 amino acid residues.A pot experiment was conducted under simulated drought stress and normal-watered conditions with three drought tolerant inbreds line "81565","N87-1","R09" and three drought intolerant line "200B","ES40","Dan340" to study.Adopting the real-time fluorescence quantitative PCR technology to detect the expression of ZmPP2Ca gene in the different maize inhreds under drought stress,the result showed that down-regulated expression was confirmed in the 3 drought-tolerant lines(81565,N87-1 and R09) and decreased to the lowest at 6 h of drought treatment.On the other hand,the 3 drought-sensitive lines(200B,ES40 and Dan340) displayed up-regulated expression under drought stress.Referring to the important role of protein phosphokinase,gene ZmPP2Ca, encoding protein phosphatase catalyzing the reverse reaction that phosphokinase catalyzes, was believed to be related with the respnose of maize to drought stress.On the other side,under the same conditional drought stress,the detect results of the activity changes of serine / threonine protein phosphatase 2 C(PP2C) in leaves showed that the PP2C activity decreased in the leaves of three drought tolerant lines with the drought treatment time;the PP2C activity in the leaves of drought intolerant lines "200B","ES40" rose after the first lower trend with the drought treatment time;the PP2C activity in the leaves of drought intolerant line "Dan 340" increased slightly overall under drought stress.These experiments suggest that the differential expression of gene ZmPP2Ca and difference of PP2C activity are speculated to be implicated in signal transduction in maize cells under drought stress.More detail researches should be conducted on the relationships between sequence of this gene and its influence to its differential expression and enzyme activity under drought stress,to provide information for development of intragenic molecular selection markers related to drought tolerance.
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