| Short-stocked and close-planting is a general development trend of modern fruit industry. The principal pathway to achieve this object is using dwarf varieties and dwarf rootstock.'Zhongai 1'is the first Chinese pear rootstock cultivar with independent intellectual property rights released by Research Institute of Pomology, Chinese Academy of Agricultural Sciences. It can result in the efficient dwarf of pear varieties, early fruiting, early full-production and high fruit quality by grafting. It can cause dwarfing and compacting and being suited as inter-rootstock for good affinity.'Zhongai 1'was popularized and widely cultured because of so many advantages. However, its molecular mechanism has not been discovered clearly by scientists. Previous studies on dwarf mechanism showed that content of GA of'Zhongai 1'were lower than other varieties in early stage. It was found that dwarfing plants were related to endogenous gibberellin auxin, other hormones and their interactions among them. Therefore, the GA maybe influences'Zhongai 1'on its height. In this study, GA20-oxidase and kaurene oxidase genes about gibberellin biosynthesis were isolated from the young leaves of'Zhongai 1'by reverse transcription polymerase chain reaction(RT-PCR)and rapid-amplification of cDNA end(RACE), and its different expression between'Zhongai 1'and plants, and provide the theoretical basis in revealing the mechanism of dwarfing'Zhongai 1'.1. Cloning GA20-oxidase and kaurene oxidase genes full-length cDNA sequences from'Zhongai 1', GenBank accession numbers were HQ833589 and JN106464, respectively. The results showed that the GA20-oxidase gene was 1314 bp, contained a length of 1179 bp complete open reading frame which encoding a protein of 392 amino acids residue. Sequence alignment results showed that the amino acid sequences had a similarity between 50% -91.58% of the amino acids of apple and other plants; kaurene oxidase gene was 1644 bp, contained a length of 1545 bp complete open reading frame which encoding a protein of 514 amino acids residue. Sequence alignment results showed that the amino acid sequences had a similarity between 77%-99% of the amino acids of apple and other plants.2. According to GA20-oxidase and kaurene oxidase genes sequence to design amplification of full-length specific primers, and amplified in'Zaosu'and'Jinxiang'to obtain their full-length cDNA sequence of GA20-oxidase and ent-kaurene oxidase genes. Sequence alignment results showed that there were only some differences nucleotide bases in the two genes among the three varieties, however, their amino acid sequence were identical. These results showed that gene function of the two genes of these three varieties were exactly same.3. GA20-oxidase and kaurene oxidase genes expression were analyzed with real time RT-PCR in different shoot growth stages in'zhongai 1','Zaosu'( as control) and'Jinxiang'(female). The resules indicated that:The expressions of GA20-oxidase gene were different in various shoot growth stages, expression content of'Zhongai 1'was lowest, secondly was"Zaosu', expression content of'Jinxiang'was largest. The expression intensity of GA20-oxidase gene in leaves of the three pear varieties had the same trend as'Zhongai 1'<'Jinxiang'<'Zaosu'in all tested growing periods. Expressions in different shoot growth stages were slightly different, in early shoot growth stage,'Zaosu'and"Jinxing'appear a expression peak at the same time, there was no expression peak in"Zhongai 1'; there was no obvious fluctuate in other shoot growth stages. Expression s of ent-kaurene oxidase gene in early shoot growth stage in'Zaosu'significantly larger than in'Jinxing'and'Zhongai 1', there was no obviously difference in other shoot growth stages in three varieties. From above results we concluded that'Zhongai 1'which dwarfing might be related to lower expression of GA20-oxidase gene.
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