摘要
采用MADS(MCM1-Agamous-Deficiens-SRF)盒基因家族功能区保守序列PCR引物,将水稻(Oryzasativa L.ssp indica)“珍汕97B”悬浮细胞、愈伤组织、分化愈伤组织和再生试管苗等不同形态发生的组织的mRNA反转录后选择性放大,经测序胶分离鉴定出一组差异表达的cDNA。对命名为RM1 cDNA的5＇端序列测定表明,RM1与典型的MADS基因——拟南芥agamous蛋白保守区一级结构同源性达63％,模拟二级结构相似性显著,初步确认RM1属于MADS基因家族成员。分子杂交证实,RM1在悬浮培养细胞中不表达,而在愈伤组织、分化愈伤组织和再生试管苗中活跃表达。
Total RNAs from the suspension cell line, callus, redifferentiatied callus and regenerated plantlets of rice ( Oryza saliva L. ssp. indica) 'Zhenshan 97B' were reverse-transcribed with poly (T) primers. The cDNAs were amplified by a pair of poly (T) primers and MADS (MCMl-Ag-amous-Deficiens-SRF) domain-specific primers and then separated on a sequencing gel. Several differentially expressed cDNA fragments were isolated. Sequencing of RM1, one of the differentially expressed gene cDNA fragments, showed that it contained a MADS domain, the primary structure of which shared a 63 % homology with the typical MADS-box agamous protein of Arabidopsis thaliana . Computer analysis also indicated that the secondary structure of RMl peptide was significantly similar to that of agamous protein. It was shown that RMl belonged to the MADS-box gene family. Molecular hybridization showed that RMl was silent in the suspension cells, while active in the callus , redifferentiated callus and regenerated plantlets.
