| Polyploidization is an important driving force for plant evolution. There are two main forms for polyploidy, namely allopolyploidy and autopolyploidy. Increasing genome dosages in autopolyploids often affect gene expression deeply and then affect the protein expression, eventually produce different phenotypes. Cultivate rice mainly is diploid. The autotetraploid can be gained by doubling choromosome artifically. Compared with diploid, autotetraploid has strong growth potential, bigger seed, thicker stem diameter, longer and wider leaf, stronger ability of resisting insects, diseases, drought and higher biomass than those of diploid. The autotetraploid colud especially improve higher yield because of doubling genome. According to expression of genes involved in seed size in the tetraploid rice, and analyzed the regulation mechanism and law of gene expression, this may provide the proofs for molecular breeding.In this experiment, there are four typical important rice varieties:Balilla, Nanjing 11, 93-11 and Haitian, we analyzed the the expression of genes (GW2, GW5, HGW, qGW8, GS3 and GS5) involved in seed size regulated..The main results are as follows:1.Agronomic traits related to seed development were investigated in the diploid and the corresponding autotetraploid plant. The results showed significan variation between the diploid and autotetraploid, such as, the stem thickness, flag leaf length, sword leaf width, and leaf color were apparently higher for the tetraploid rice than those of the corresponding diploid rice; however, the tillering ability became weaker. The PH of the autotetraploid 9311 and Balilla were slightly higher than their corresponding diploids; the difference in height was about 5-10 cm. Among all the traits, grain length (GL), grain width (GW), length to width (L/W) (except Nanjing 11), grain size (GS), and grain circumference (GC) showed significant (P< 0.01) variation between the diploid and autotetraploid. The KGW of autotetraploid have 31%-35% bigger than diploid, grain length increased 2-3 cm, grain width increaseed 0.5 cm.2. Based on scanning electron microscopic of young ear secondary branch formation stage and pistal and stamen formation stage of Balilla and DBalilla, we found that young ear development process of tetraploid rices was consistent with the corresponding diploid, and ensured the accuracy of the materials.3. The expression level analysis of six main seed size-related genes through the Q-PCR technology. The results are as follows:1). in 4 n rice, the positive regulator expressed increased, the negative regulator expressed decreased. This displayed that the expression results in molecular levels were accordance with the seed size phenotype.2). the expression of some genes was materials-specific. For example, GW5 were not expressed in these four rice varieties, qGW8 in Balilla was silence in early development stage, while its expression increased along with the development stages in 93-11.3). most of the gene expression changes accorded with seed development process, belong to stage-specific. For example, at the beginning of young ear development in 93-11, most of genes, except GS3 and qGW8, the expression levels were increased in stage 3 and 4, and were the highest for express levels, but then begun to decline.4). There was different effect gene in different development period. In stage 1, the negative regulatory factors GW2, GS3 were expressed. However, they presented tendency declined in 4n rice 93-11, whlie in Dbalilla, positive factors HGW, GS5 and qGW8 expressed significantly higher than that in 2n rice; In stage 3, GS3 expressed the highest in 4 n rice 93-11 as a major gene; In stage 5, qGW8 expressed the highest in 4 n rice 93-11 as a major gene. These results showed that different major gene played an important role in different development period.4. We verified that the over-expresseing gene regulate seed size by transgenic ex periments. In Balilla (2n), we overexpressed the negative regulator GW2, and got 2 8 transgenic plants. We detected the markers of these transgenic plants (hyg), and a bout 800 bp target fragment could be amplified, proved that the gene has been tran sferred to plant. The detection of the expression differences by Q-PCR showed that the gene expression level was orded:Balilla (2n)T> Balilla (2n)> Balilla (4n) (1 28.0>54.7> 17.44). And corresponding plant seed size traits was orded:BalillaT<BaIi lla (2n)< Balilla(4n) (grain width:2.95cm<3.61cm<3.73cm,1000-grain weight:22.3g< 31.82g<49.41g). The results showed that dose changes of the negative regulator GW 2 were consistent with its function.In short, these studies suggest that the gene expressions involved in seed size are associated with rice ploidy, development stages and the gene function. It perhaps implys that we may regulate the expression of individual major gene to produce bigger seed for diploid rice. Besides, this study provids new resources or way for rice breeding. |
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