As a base for human transcriptome and functional genomics, we created the "full-length long Japan" (FLJ) collection of sequenced human cDNAs. We determined the entire sequence of 21,243 selected clones and found that 14,490 cDNAs (10,897 clusters) were unique to the FLJ collection. About half of them (5,416) seemed to be protein-coding. Of those, 1,999 clusters had not been predicted by computational methods. The distribution of GC content of nonpredicted cDNAs had a peak at ∼58% compared with a peak at ∼42%for predicted cDNAs. Thus, there seems to be a slight bias against GC-rich transcripts in current gene prediction procedures. The rest of the cDNAs unique to the FLJ collection (5,481) contained no obvious open reading frames (ORFs) and thus are candidate noncoding RNAs. About one-fourth of them (1,378) showed a clear pattern of splicing. The distribution of GC content of noncoding cDNAs was narrow and had a peak at ∼42%, relatively low compared with that of protein-coding cDNAs.

By analyzing 1,780,295 5Ј-end sequences of human full-length cDNAs derived from 164 kinds of oligo-cap cDNA libraries, we identified 269,774 independent positions of transcriptional start sites (TSSs) for 14,628 human RefSeq genes. These TSSs were clustered into 30,964 clusters that were separated from each other by more than 500 bp and thus are very likely to constitute mutually distinct alternative promoters. To our surprise, at least 7674 (52%) human RefSeq genes were subject to regulation by putative alternative promoters (PAPs). On average, there were 3.1 PAPs per gene, with the composition of one CpG-island-containing promoter per 2.6 CpG-less promoters. In 17% of the PAP-containing loci, tissue-specific use of the PAPs was observed. The richest tissue sources of the tissue-specific PAPs were testis and brain. It was also intriguing that the PAP-containing promoters were enriched in the genes encoding signal transduction-related proteins and were rarer in the genes encoding extracellular proteins, possibly reflecting the varied functional requirement for and the restricted expression of those categories of genes, respectively. The patterns of the first exons were highly diverse as well. On average, there were 7.7 different splicing types of first exons per locus partly produced by the PAPs, suggesting that a wide variety of transcripts can be achieved by this mechanism. Our findings suggest that use of alternate promoters and consequent alternative use of first exons should play a pivotal role in generating the complexity required for the highly elaborated molecular systems in humans.[Supplemental material is available online at www.genome.org. The sequence data from this study have been submitted to DDBJ under accession nos. DA000001-DA999999, DB000001-DB294747, DB294748-DB384947, BP192706-BP383670, AU279383-AU280837, and AU116788-U160826.]One of the most striking findings revealed by the Human Genome Project is that the human genome contains only 20,000-25,000 kinds of protein-coding genes (International Human Genome Sequencing Consortium 2004). This number is unexpectedly small compared with the total gene numbers in yeast, fly, and worm genomes, which are estimated to be 6,000, 14,000, and 19,000, respectively (Goffeau et al. 1996;C. elegans Sequencing Consortium 1998;Adams et al. 2000). It is supposed that there must be other factors in addition to mere gene numbers to satisfy the prerequisites that enable the human genome to fabricate such highly elaborated systems as the brain and immune systems. To explain this, it has been hypothesized that multifaceted use of the genes should play a pivotal role in functional

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Treatment with 5-azacytidine, a DNA demethylating reagent, induced flowering in Perilla frutescens (L.) Britton var. crispa (Thunb. ex Murray) Decne. ex L. H. Bailey, an absolute short-day plant under long days. The 5-azacytidine treatment induced slight suppression of vegetative growth but had no obvious effect on any other phenotypes. The Southern hybridization analysis of the genomic DNA isolated from the leaves of 5-azacytidine-treated plants and digested with restriction enzyme, methylation-insensitive Msp I or methylation-sensitive Hpa II with P. frutescens 25S-18S rDNA intergenic spacer probe indicated that the 5-azacytidine treatment caused demethylation of the genomic DNA. The 5-azacytidine-induced flowering was delayed as compared with the short day-induced flowering. Flowers were formed even at the lower nodes which had not been directly treated with 5-azacytidine. The results suggest that DNA demethylation induced flowering by inducing the production of a transmissible flowering stimulus in P. frutescens.

To investigate the effects of plant species in grassland on methane (CH4) and nitrous oxide (N2O) fluxes from soil, fluxes from an orchardgrass (Dactylis glomerata L.) grassland, white clover (Trifolium repens L.) grassland and orchardgrass/white clover mixed grassland were measured weekly from April 2001 to March 2002 using a vented closed chamber method. Related environmental parameters (soil inorganic N content, soil pH (H2O) value, soil moisture content, soil temperature, grass yield, and the number of soil microorganisms) were also regularly monitored. On an annual basis, CH4 consumption in the soil of the orchardgrass grassland, white clover grassland and orchardgrass/white clover mixed grassland was 1.8, 2.4, and 1.8 kg C ha−1 year−1, respectively. The soil bulk density of the white clover grassland was lower than that of the other grasslands. Fluxes of CH4 were positively correlated with the soil moisture content. White clover increased the CH4 consumption by improving soil aeration. Nitrogen supply to the soil by white clover did not decrease the CH4 consumption in the soil of our grasslands. On the other hand, annual N2O emissions from the orchardgrass grassland, white clover grassland, and orchardgrass/white clover mixed grassland were 0.39, 1.59, and 0.67 kg N ha−1 year−1, respectively. Fluxes of N2O were correlated with the NO3− content in soil and soil temperature. White clover increased the N2O emissions by increasing the inorganic N content derived from degrading white clover in soil in summer.