| In this study regeneration system of apple seedlling was developed, Agrobacterium-mediated transformation was adopted to introduce the LEAFY gene into apple seedlling, and new germplasm with LEAFY gene was obtained, which was the basic of researching the flowering mechanism of apple. The detailed results were listed as follows:1. Effecting factors of expanding propagation and regeneration of hybrid of apple was studied, including type and content of hormone, time of darkness treatment, ways of leaf deposition. The results showed that expanding propagation medium was MS+NAA0.1mg/L+BA0.5mg/L, rooting medium was l/2MS+IBA1.0mg/L and darkness treatment 7-14 days, regeneration medium was MS+NAA0.4mg/L+TDZ2.0mg/L and darkness treatment 21 days, while adxial surface touch medium. Regenerationg rate was 0.5%.2. Factors that influence percentage of regeneration including instension of selection, dipping time, the time of co-culture, content of antibiotic were studied. The effective transformation system of hybrid of apple was developed as follow: Leaves were dipped into Agrobecterium suspension added 150mg/L AS for five minutes, then were co-culture on regeneration medium of hybrid of apple for 2 days in darkness , then were co-culture with cef 300mg/L and kml5mg/L in light, could get the highest 41.4% gus transient expression.3. The GUS histochemistry assay and PCR,RT-PCR Amplification on these resistant plants approved that LEAFY gene has been integrated into the host genome of hybrid of apple in two plants and the gene has expressed.4. Proteins isolated by two-dimensional gel electrophoresis in transformation plants and normal plants was 207 proteins which matched, PDQuest software analysis showed that the matching rate was 92%. There were 12 distinct proteins in transformation plants, among which 4 proteins were added and 6 proteins were up-regulated and 2 proteins was lost.
|
PDF Full Text Request