A suppression subtractive hybridization library construction for red skin mutant of ‘Zaosu’ pear and MYBR gene analysis
2012
Liu, YJ | Du, YM | Xu, LF | Hu, M | Li, ZH
Red pear breeding is one of the major domestic and international pear breeding projects. The aim of this study is to investigate red skin development-related genes of the red skin mutant of ‘Zaosu’ (Pyrus bretschneideri Rehd.) pear and provide important information for elucidating the molecular mechanism of red pigment formation in the skin. A cDNA library was constructed using the suppression subtractive hybridization (SSH) method, in which the green-skinned pear ‘Zaosu’ served as the driver, and its red skin mutant ‘Zaosu Red’ served as the tester. All positive clones between 250–1000 bp were chosen and sequenced from the library, and 735 expressed sequence tags (ESTs) were obtained. According to BLAST screening and functional annotation in the GenBank database, 314 unique gene fragments were obtained. Among the obtained fragments, 198 ESTs were found to have homologous genes with known functions, 61 ESTs with unknown functions and 55 ESTs with no homologous genes. Results showed that the differentially expressed genes are involved in anthocyanin biosynthesis, signal transduction, energy metabolism, transcription, disease resistance and bud mutation. Moreover, a novel cDNA of ‘Zaosu Red’ MYBR gene was isolated with the strategy of in silico cloning in combination with RT-PCR from ‘Zaosu Red’ pear fruit. The full-length of MYBR cDNA was 1552 bp, with an open reading frame from 12 to 1355 bp, coded 447 amino acids.
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