| Sugar beet is a important sugar crop in world. It is difficult to breed the varieties with both high yield and high sugar content due to yield has the negative correlation with sugar content within sugar beet. Some researches have shown that sucrose phosphate synthesis contributed much in the distribution of carbohydrates.During recent years, genetic modification has made large progress in crop quality improvement Because of its high efficiency, easy control, short time consume, low cost, and far relative gene resources also can be used, so genetic modification has been a efficient approach of breeding new crop varieties. Cloning target genes from high quality crop and transformation into new crops modified have been exploited. It has been a developmental direction of modem molecular breeding to obtain new high quality crop varieties (lines).In sugar beet genetic transformation, the efficiency was very low. It is difficult to establish stable regeneration system of sugar beet plants. In this study, maize SPS gene has been cloned, constructed recombined plasmid vector. Using Agrobaterium-mediated transformation and particle bombardment transformation, SPS gene has been introduced into sugar beet plants. The factors of influencing plant regeneration and transformation efficiency have been studied. This study established stable transformation system and plant regeneration system. Also acquired sugar beet transgenic plants which integrated maize SPS gene. The main results as follows:1. The whole length maize SPS gene has been cloned from maize total RNA through RT-PCR. The fragment has a full length open reading frame of 3200 base pairs encoding 1066 amino acids. The gene has 99% homology with SPS gene nucleic acid sequence reported previously.2. In PCR amplification, the real Taq polymerase has highest performance when concentration of Mgcl2 was 0.2 mM.3. Two transformation vectors have been constructed, one was PBLSPS, another one was PBSPS. PBLSPS was designed for Agrobaterium-mediated transformation, while PESPS was used to particle bombardment transformation, which has no selection marker.4. This research has established organogenesis regeneration system of sugar beet petiole.a The best disinfectants combination to sugar beet seeds was absolute sulphate, 70% ethanol, 3% NaCIO, with 0.1% Hgcl2.b. Best culture medium for shoots inducing was MS+IBA 0.1mg/l+ BA1.0mg/l+30g/l sucrose +9g/l agar, PH5.8.c. The most appropriate explant for shoots inducing was petioles pre-cultured by 30 days.d. 4 sugar beet lines has been proved with high shoots inducing frequency from 12 sugar beet lines. That was, shoots inducing frequency of 8910A and 8908B were 51.6%, while 8910B and Xinshuang 8-3-2 were 48.3%.e. Petioles from leaf shoots have been firstly used as explant to induce adventitious shoots.5. Transformation system of sugar beet petiole pre-cultured has been optimized in Agrvbaterium-mediated transformation.a When Agrobaterium concentration was 0.8 OD600 and infectious time was 8 min., sugar beet lines 8910A and 8908B can reach high shoots inducing frequency, which were 36.6% and 33.3% respectively. Comparing line Xinshuang 8-3-2, shoots inducing frequency was 40% when Agrobateriwn concentration was 0.1 OD600 by 8 min infectious.b. The appropriate culture temperature of sugar beet was 22-25℃.c. The concentration of kanamycin for shoots selection was 150mg/l. During rooting period, 20mg/l of its concentration has been identified to be suitable for lines 8910B and 8908B. While for line Xinshuang 8-3-2, it should be 25mg/l.d. Late selection system approach has been used during shoots inducing period. Transgenic explants were selected both at the stage of differentiation and root regeneration of shoots.6. For the first time, this study acquired transgenic sugar beets integrated with maize SPS gene trough Agrobaterium-mediated transformation. 4 transgenic plants has been identified by PCR and Southern blotting, the percentage of p...
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