CHARACTERIZATION OF GENE REPERTOIRES AT MATURE STAGE OF CITRUS-FRUITSTHROUGH RANDOM SEQUENCING AND ANALYSIS OF REDUNDANT METALLOTHIONEIN-LIKE GENES EXPRESSED DURING FRUIT-DEVELOPMENT

We carried out a random sequencing of cDNA library derived from mature citrus fruit (Citrus unshiu Marc.) for identifying the gene repertoir es expressed at the mature stage. Among 297 clones analyzed, 195 cDNA clones (65.7%) were putatively identified to previously characterized genes with optimized (OPT) scores of greater than or equal to 200 thro ugh a homology search to DNA database, whereas 102 clones (34.3%) resu lted in low OPT scores (<200) and did not show any significant sequenc e identity with previously published genes. Among them, clones homolog ous to metallothionein (MT)-like genes appeared 62 times, being mostly redundant, and accounting for about 20.9% of the total 297 clones. To gain a better understanding of the MT-like genes, two types of cDNA c lones were isolated. One clone (CitMT36) resembled the type 2 MT gene containing Cys-X-Cys motifs in both N- and C-terminal, but the consens us sequence in the N-terminal domain, Cys-Cls and Cys-X-X-Cys was modi fied in CitMT36 to X-Cys and Cys-X-X-X, respectively. We suggest that these form a 'novel type 2' group of MT-like clones. The other clone ( CitMT45) showed homology to type 3 MT-like genes, which have been foun d in mostly fruit tissues so far. By Southern blot analysis, both clon es showed one or two bands, suggesting that both CitMT36 and CitMT45 a re present in single or a few copies in the citrus genome. Transcripts of CitMT36 were evenly detected in all tissues examined, whereas thos e of CitMT45 were detected primarily in fruit during the developmental phase. Neither of the MT-like genes was induced in leaves by Zn and C u. Collectively, MT-like genes from citrus would be regulated differen tially depending on the fruit developmental stage and organs, indicati ng a change in their expression under the different physiological and molecular environment of fruit cells. (C) 1998 Elsevier Science B.V. A ll rights reserved.