Phylogenetic Relationships of Pomegranate Accessions by Using rDNA ITS, RAPD and ISSR Molecular Markers

• Main Authors: Wan-Ling Wang, 王婉伶
• Other Authors: Jwu-Guh Tsay
• Format: Others
• Language: zh-TW
• Published: 2004
• Online Access: http://ndltd.ncl.edu.tw/handle/32408601349252427847

碩士 === 國立嘉義大學 === 園藝學系研究所 === 94 === In order to establish pomegranate cultivation in Taiwan, selected pomegranate lines which were introduced from Mainland China, Singapore and United States, had been assayed for their growth and physiological characteristics, phylogenetic relation by contents of polyphenolic compounds, ITS sequence of rDNA, ISSR and RAPD markers. The investigation was performed using four large-fruited lines of three-year-old plants. Gradual increase of the fresh fruit weight, width, length, and volume was observed during the experimental period. Twenty days post-fruit setting, fruits grew more in width than in length. Thirty days post-fruit setting, its pericarp weight was greater than seed weight. However, fifty days post-fruit setting, the seed weight was greater than the pericarp weight. The result showed that four pomegranate lines with different shapes were affected mainly by genetic components. On the other hand, soluble solid contents increased but accompanied decrease in acidity during fruit development. At hundred days post-fruit setting, soluble solid juice contents of A-01 and B-02 lines were reached the maximum. For the period of maximal full bloom, A-01 and A-02 lines flowered approximately at September whereas B-01 and B-02 lines reached in December. The analysis of fruit setting percentage, quality, yield and full bloom suggests that pomegranate will be harvested for marketing and breeding at hundred day post-fruit setting. Polyphenol contents of pericarp from 31.8% to 45.6% were measured by HPLC. B-01 line had the highest content among 4 Singapore lines. Based on cluster analysis of polyphenol data, five pomegranate lines could be divided into two groups: one group included A-02, B-01 and B-02, the others were A-01 and the commercial fruit line. To use the molecular technology to differential these accessions, sequence analysis of 5.8S rDNA and internal transcribed spacer (ITS) in eight samples of five lines. It showed no differences among 5 lines. In RAPD analysis twenty-four polymorphic bands were amplified by six primers, The polymorphism rate was 28.37%. A clone-specific band was only found in Singapore lines. Using UPGMA clustering analysis of RAPD polymorphic bands, the pomegranate could be divided into five groups with their similarities of 71%. Although many polymorphic bands were produced amongst different lines none showed any correlation with fruit color or growth characters. The cophenetic correlation coefficient of Nei/Li's and Jaccard's similarity coefficient of RAPD was 0.995. Dinucleotides motif: (GA)n sequence presented major variations in pomegranate genome through ISSR analysis. Based on UPGMA clustering analysis of ISSR polymorphic bands, the pomegranate could be divided into three groups with their similarities of 87% to 88%. Pomegranate lines of 3 origins could be distinguished by seven polymorphic bands. The result of ISSR clustering analysis coincided with the origins of pomegranate lines. The cophenetic correlation coefficients of RAPD and ISSR were 0.467 and 0.351, respectively. It suggested that two molecular markers could be used in identifying part of pomegranate lines. Key words: Pomegranate; Flowering; Fruit growth and development; polyphenolic compounds; ITS; RAPD; ISSR; Similarity; Identification