GENE TRANSFORMATION EXPERIMENT OF THE SELF-INCOMPATIBILITY RELATED GENES

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Objectives : To understand the molecular mechanism of self-incompatibility and to assess a possibility of molecular breeding of this character, molecular characterization and gene manipulation was conducted on the S-related genes in sporophytic self-incompatible plant : Brassica campestris and in gemetophytic self-incompatible plant : Pyrus pilifolia.Results : In B.campestris, SLG8, SLG9, SLG12, SRK8, SRK9, SRK12, SRA1, SRA2, SRA3, SRA4 and SRBs were isolated from cDNA or genomic DNA libraries and sequenced. Dendrograms constructed on the basis of synonymous and nonsynonymous substitutions indicated that SRA differentiated first, followed by SRB and SLG/SRK.When the SLG8 genomic clone was introduced into B.campestris some of transformed plants changed to self-compatible. Furthermore, the antisense SLG gene was transformed into the same species. Analysis of a transformant showed that the self-incompatibility was broken down by the action of the introduced antisense gene. These results i … More ndicated the self-incompatibility could be controlled, at least partly, by gene manipulation. The genes encoding 3 RNases were cloned from the style of a self-incompatible cultivar, Nijisseiki (S2S4), and its self-compatible mutant, Osa-Nijisseiki (S2S4sm, sm means stylar part mutant), of Japanese pear. For Nijisseiki, cDNAs coding for two S-RNases (S2-RNase and S4-RNase) and an RNase unrelated to self-incompatibility (non-S-RNase) were cloned from the stylar cDNA library. However, the cDNA coding for S4-RNase was neither amplified by PCR nor cloned from the library of Osa-Nijisseiki, suggesting that the mutation of self-incompatible Nijisseiki to self-compatible Osa-Nijisseiki is due to a failure of expression of S4-RNase. The S3-gene promoter of Petunia inflata was used in constructing the vectors for the Japanese pear S-genes. The binary vectors of the Agrobacterium were transformed in the petunia resulting in shoot regeneration. In vitro cultures of the Japanese pear were also extended. Less