Rapid cloning and bioinformatic analysis of spinach Y chromosome-specific EST sequences

Journal of genetics

Long-Dou Lu¹  Wei-Li Zhang¹  Shao-Jing Wang¹  Shu-Fen Li¹  Chuan-Liang Deng1¹  Wu-Jun Gao¹  Ying Cao¹ 

[1] College of Life Science, Henan Normal University, Xinxiang 453007, People’s Republic of China$$

关键词: spinach;    Y chromosome;    microdissection;    hybridization-specific amplification;    expressed sequence tag.;   

DOI  :  

学科分类：生物科学（综合）

来源: Indian Academy of Sciences

【 摘 要 】

The genome of spinach single chromosome complement is about 1000 Mbp, which is the model material to study the molecular mechanisms of plant sex differentiation. The cytological study showed that the biggest spinach chromosome (chromosome 1) was taken as spinach sex chromosome. It had three alleles of sex-related 𝑋, 𝑋m and 𝑌. Many researchers have been trying to clone the sex-determining genes and investigated the molecular mechanism of spinach sex differentiation. However, there are no successful cloned reports about these genes. A new technology combining chromosome microdissection with hybridization-specific amplification (HSA) was adopted. The spinach Y chromosome degenerate oligonucleotide primed-PCR (DOP-PCR) products were hybridized with cDNA of the male spinach flowers in florescence. The female spinach genome was taken as blocker and cDNA library specifically expressed in Y chromosome was constructed. Moreover, expressed sequence tag (EST) sequences in cDNA library were cloned, sequenced and bioinformatics was analysed. There were 63 valid EST sequences obtained in this study. The fragment size was between 53 and 486 bp. BLASTn homologous alignment indicated that 12 EST sequences had homologous sequences of nucleic acids, the rest were new sequences. BLASTx homologous alignment indicated that 16 EST sequences had homologous protein-encoding nucleic acid sequence. The spinach Y chromosome-specific EST sequences laid the foundation for cloning the functional genes, specifically expressed in spinach Y chromosome. Meanwhile, the establishment of the technology system in the research provided a reference for rapid cloning of other biological sex chromosome-specific EST sequences.

【 授权许可】

Unknown