Morphological Characteristics, Expression And Function Analysis Of A New Allelic Mutant Of DROOPING LEAF(DL) Gene

DROOPING LEAF (DL) gene is a member of YABBY gene family, and is an ortholog of the CRABS CLAW (CRC) gene of Arabidopsis thaliana. It controls the development of carpel in the flower and the formation of midrib in the leaf. Loss-of-function mutations of DL present midrib-less drooping leaves and degrees of abnormalities in whorl 4 of the flower. We found a new allelic mutant of DL gene, called dl-5. In this study, we did further research about the function of DL via analysing the morphological characteristics, expression and protein structure of dl-5. The consequences are as follows:dl-5 showed a quite different phenotype from the other mutants found before. The DL/dl-5 heterozygote exhibited defect of midrib in its upper leaves while appeared normal in the lower leaves, and showed lower fertility. In addition, unlike the other severe mutants which showed homologous transformation of carpels into stamens, dl-5 developed several whorls of leafy organs at the position of the gynoecium. These organs were coaxial and arranged in a alternate pattern. As a consequence, dl-5 showed a quite different phenotype from the other mutants of DL gene, and was a semi-dominant allele.Using map-based cloning, we mapped dl-5 between the markers RM14591 and SSR1 on the short arm of rice chromosome 3. The physical distance was 21kb, in which DL was the only candidate gene. Sequence analysis revealed that a single base change from G to T caused an amino acid replacement from Cys (42) to Phe in the second extron. Protein structure analysis showed that the mutated site was one of the presumptive key sites in the Zinc-finger domain.We examined the expression level of DL and some MADS-box genes which were related to floral development in the gynoecium using real-time PCR, and the results showed that class C and D genes and class E gene OsMADS1 were down-regulated significantly; other class E genes and B genes changed slightly. The class B gene OsMADS16 which interacts antagonistically with DL gene was not up-regulated, which probably because the expression of DL gene was enough to suppress OsMADS16 though down-regulated. This must be the main reason why dl-5 didn’t appear the transformation of pistil to stamen.