| The heading process is important and unique for the Chinese cabbage (Brassica rapa L.ssp. pekinensis). Expressed sequence tag (EST) analysis would result in a gene expression profile of any unique biological processes. To understand the mechanism of Chinese cabbage heading process, we randomly sequenced over 1000 ESTs from the cDNA library constructed before at our laboratory from Chinese cabbage leaves in the early phase of the heading stage. In order to get the recombinant plasmid DNAs for EST analysis, our first step was to infect Escherichia coli DH10B with the Lambda ZipLox cDNA library, and 2697 plasmid clones probably carrying inserted cDNA were then obtained through in vivo excision. We randomly selected 1361 plasmid clones, extracted their plasmid DNAs and sequenced them with our DNA sequencer (Beckman & Coulter CEQ2000). The sequencing results indicated that 127 clones had inserts shorter than 150bp or only polyA sequences and 72 clones had no cDNA inserts, and the remaining 1162 clones were used for further analysis.The SeqMan II module of DNAStar software was used for contig analysis of all the 1162 ESTs, and a total of 902 different contigs were obtained. 749 of the 902 contigs were composed of only one EST, and such contig was also called singleton. This result suggested that there were many different genes expressed during the heading process of Chinese cabbage.The 1162 ESTs were edited manually and then compared with the sequences in the non-redundant protein and nucleotide databases of NCBI, 902 ESTs sharing significant similarity with sequences registered in these databases after being analyzed using BLASTX and BLASTN program. Out of these, 670 ESTs might encode 471 genes of known function and the other might represent 200 hypothetical proteins of unidentified function. Sixty of the remaining 260 ESTs had no homology with any of the registered EST sequences currently released from the GenBank after being aligned with our local 'est_others' database. These putatively new ESTs were very important for understanding the unique development process and molecular genetic mapping in Chinese cabbage.The results of a series of BLAST searches suggested that 94.8% (1102/1162) ESTs had homologues at the protein or nucleotide level, which was not reported at the research of the same plant species. By analyzing the sources of the organisms, it was found that about 73.6% of the functionally known protein genes were from Arabidopsis thaliana; however, at the nucleotide level, only 51% (592/1162) of the total ESTs showed highest homology with those from Arabidopsis thaliana.These results suggested that gene expression pattern in Chinese cabbage was different from that in Arabidopsis thaliana.The functional assignment of ESTs in Chinese cabbage was carried out based on the method used in the Arabidopsis thaliana genome sequencing initiative. The results indicated that the ESTs and the genes involved in the process of protein synthesis had the highest proportion innot only the numbers but also the types of expressed genes in the heading process of Chinese cabbage, and the ESTs encoding the proteins and enzymes in the process of energy metabolism, cell defense, cell structure, organization and biogenesis, transcription, protein destination and storage protein and signal transduction were all present in multiple copies. When compared the expression profile of the heading leaves of Chinese cabbage with that of other dicotyledonous plant leaves, much difference was found in the expression of the three functional categories of ESTs for protein synthesis, energy metabolism, and cell structure, organization and biogenesis process. We presumed that these differences were related to the special needs of the morphological development during heading process of Chinese cabbage.The expression profile analysis of the early phase at the heading stage of Chinese cabbage would benefit the understanding of the special heading process in Chinese cabbage, Brassica oleracea and other related species at the mole...
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