Studies on Somaclonal Variation Derived from the Suspension Culture of PLBs in Oncidium

• Main Authors: Chiu-Yen Pan, 潘秋燕
• Other Authors: Li-Jen Liao
• Format: Others
• Language: zh-TW
• Published: 2000
• Online Access: http://ndltd.ncl.edu.tw/handle/86223099360210397932

碩士 === 國立高雄師範大學 === 生物科學研究所 === 88 === In Taiwan Oncidium is an important cut flower which is manly exported to Japan. Major plantlets production method is mericlonal propagation with PLBs using tissue culture techniques. Plants regenerated from undifferentiated tissue cultures have become a new and useful source of genetic variation. Variation generated by the use of a tissue culture cycle has been termed somaclonal variation by Larkin and Scowcroft (1981). They defined a tissue culture cycle as a process that involves the establishment of a dedifferentiated cell or tissue culture under defined conditions, proliferation for a number of cell generations, and the subsequent regeneration of plants. Oncidium hybrid ''Grower Ramsey'' was used in this study to understand the potential and frequency of somaclonal variation derived from tissue culture procedure or period. The results obtained are summarized as follows: 1. Ochidium PLBs suspension cultured in KC liquid medium supplement with 10 μM BA had the best growth response, the other suspension culture systems supplement with 10 μM NAA, 10 μM 2,4-D, 10 μM BA+10 μM NAA or 10 μM BA+10 μM 2,4-D turned brown gradually. 2. Use the random primers OPU-09 and OPJ-09 for RAPD analysis to detect the somaclonal variation between the In vitro Oncidium basal population and PLBs suspension culture that treated with different PGR. The detectable sensitivity of these two primers for somaclonal variation are different, the OPJ-09 is better than OPU-09. But as the result of OPU-09 RAPD analysis, a lot of DNA sequences that are complementary to OPU-09 primer, have the better stability even though the PLBs suspension culture treated with different PGR long period about 9 months. 3. Oncidium protoplasts can be isolated from mesophyll tissue or PLBs. The best isolation enzyme combination was the solution contained KC mineral salts supplemented with 2% cellulase RS, 1% macerozyme R-10 and 0.5 M mannitol as osmoticum at pH 5.8. The protoplasts derived from mesophyll tissue or PLBs cultured in KC medium containing 5 μM NAA or 10 μM NAA had a best response in cell division. However, cell colonies would die 1 month after initial culture.