Construction and analysis of a suppression subtractive hybridization (SSH) library of genic multiple-allele inherited male-sterility in Chinese cabbage (Brassica campestris L. ssp. pekinensis)

Abstract

Utilization of male sterility is a key method for producing crossbred Chinese cabbage (Brassica rapa L. ssp. pekinensis (Lour.) Olsson. In this study, suppression subtractive hybridization (SSH) was used to construct sterility and fertility cDNA libraries, which included differentially, expressed clones between fertile and sterile buds of the A/B line ‘AB01’. The positive clones were randomly selected by polymerase chain reaction amplification (PCR) and 25 high quality sequences (22 from the fertile-tester library and three from the sterile-tester libraries) were generated. The fragment lengths varied from 77 to 469 bp. Differential expression patterns between fertile and sterile buds were selected and verified using five expressed sequence tags (ESTs). Results indicated that, three ESTs were expressed only in fertile buds and two ESTs were down-regulated in sterile buds. According to the Basic Local Alignment Search Tool (BLAST) screening and functional annotation, the 25 ESTs were homologous to known sequences deposited in National Center for Biotechnology Information (NCBI). These genes had homology to known proteins such as flowers/buds development proteins, metabolic-related proteins, cell structure proteins, cell growth/division proteins and secondary metabolic-related proteins. The results suggested that, these proteins played a critical role in nuclear male sterility progression of genic multiple-allele inherited male-sterility in Chinese cabbage.

Key words: Chinese cabbage, male sterility, suppression subtractive hybridization (SSH), expressed sequence tags (ESTs).