In order to provide information for the development of molecular selection markers for drought tolerance improvement, the methods of prometric analysis, quantitative real-time PCR and field evaluation were employed for the identification of the differential expression of candidate genes under drought stress in maize. At seventeen, twenty-four and forty-eight hours of polyethylene glycol-simulated drought stress at the seventh leaf stage, leaf samples were collected from two drought-tolerant inbred lines for prometric analysis by two-dimensional electrophoresis and peptide mass fingerprinting. Fifty-eight proteins out of more than 500 were found in response to drought stress. Three drought-induced spots 2506, 3507 and 4506 showed sequence similarity with cinnamyl alcohol dehydrogenase, cytochrome protein 96A8 and S-adenosyl-L-methionine synthase, respectively. The expression of two key enzymes to lignin biosynthesis was quantified by quantitative real-time PCR among three drought-tolerant and one drought-sensitive inbred lines under drought stress and well-watered control conditions. After a decrease at the beginning of drought stress, the expression of cinnamyl alcohol dehydrogenase and caffeate O-methyltransferase recovered at twenty-four hours of the drought stress in the three drought-tolerant lines, but not in the drought-sensitive lines. Leaf lignin content, anthesis-silking interval and grain weight per plant were investigated with six inbred lines of varying drought tolerance under drought stress and well-watered control. Drought tolerance coefficients of these three characters were calculated and the correlation coefficients among these drought tolerance coefficients were estimated. Significant difference in leaf lignin content was found among the inbred lines and in response to drought stress. Close correlations were observed between the drought tolerant coefficients for leaf lignin content and grain weight per plant, and between the drought tolerant coefficients for leaf lignin content and anthesis-silking interval. These results indicate that leaf lignin content is a useful index for evaluation of drought tolerance in maize. Molecular selection markers can be developed on the basis of differential expression of the candidate genes and applied to maize improvement for drought tolerance.

The in silico mapping (ISM) technique and its extension represent major advances for novel gene discovery in germplasm resources of inbred lines. However, the techniques suffer from a relatively high false-positive rate (FPR) and they do not consider the effect of linkage disequilibrium (LD) markers around the identified quantitative trait locus (QTL). In addition, it has not yet been established whether it is optimal to use absolute trait differences as the response variable. To address these problems, this article presents the multiple loci ISM (MLISM) approach, which uses all markers on the entire genome, along with a penalized maximum likelihood. The method proposed here was verified by a series of simulation experiments with a maize pedigree population of inbred lines of known ancestry. Results from the simulated studies show that the best response variable is the trait product. The MLISM FPR is substantially decreased and the proportion of the number of false QTL to the number of LD markers around the identified QTL is adequately reduced. The MLISM method, with the trait product as the response variable, is an improvement on the existing methods for novel QTL mapping in germplasm resources of inbred lines.