Characterization of expressed sequence tags from a full-length enriched cDNA library of Cryptomeria japonica male strobili

<p>Abstract</p> <p>Background</p> <p><it>Cryptomeria japonica </it>D. Don is one of the most commercially important conifers in Japan. However, the allergic disease caused by its pollen is a severe public health problem in Japan. Since large-scale analysis of expressed sequence tags (ESTs) in the male strobili of <it>C. japonica </it>should help us to clarify the overall expression of genes during the process of pollen development, we constructed a full-length enriched cDNA library that was derived from male strobili at various developmental stages.</p> <p>Results</p> <p>We obtained 36,011 expressed sequence tags (ESTs) from either one or both ends of 19,437 clones derived from the cDNA library of <it>C. japonica </it>male strobili at various developmental stages. The 19,437 cDNA clones corresponded to 10,463 transcripts. Approximately 80% of the transcripts resembled ESTs from <it>Pinus </it>and <it>Picea</it>, while approximately 75% had homologs in <it>Arabidopsis</it>. An analysis of homologies between ESTs from <it>C. japonica </it>male strobili and known pollen allergens in the Allergome Database revealed that products of 180 transcripts exhibited significant homology. Approximately 2% of the transcripts appeared to encode transcription factors. We identified twelve genes for MADS-box proteins among these transcription factors. The twelve MADS-box genes were classified as <it>DEF/GLO/GGM13-, AG-, AGL6-, TM3- </it>and <it>TM8</it>-like MIKC^Cgenes and type I MADS-box genes.</p> <p>Conclusion</p> <p>Our full-length enriched cDNA library derived from <it>C. japonica </it>male strobili provides information on expression of genes during the development of male reproductive organs. We provided potential allergens in <it>C. japonica</it>. We also provided new information about transcription factors including MADS-box genes expressed in male strobili of <it>C. japonica</it>. Large-scale gene discovery using full-length cDNAs is a valuable tool for studies of gymnosperm species.</p

ヤナギ属植物の環境適応性と雌雄性に関する分子生物学的研究

University of Tokyo (東京大学

Comprehensive analysis of small RNAs expressed in developing male strobili of <i>Cryptomeria japonica</i>

<div><p>Deep sequencing of small RNAs (sRNAs) in developing male strobili of second-generation offspring originating from a nuclear genic male sterile tree of <i>Cryptomeria japonica</i> were performed to characterize sRNA populations in the male strobili at early pollen developmental stages. Comparing to sequences of microRNA (miRNA) families of plant species and sRNAs expressed in the reproductive organs of representative vascular plants, 37 conserved miRNA families were detected, of which eight were ubiquitously expressed in the reproductive organs of land plant species. In contrast, miR1083 was common in male reproductive organs of gymnosperm species but absent in angiosperm species. In addition to conserved miRNAs, 199 novel miRNAs candidates were predicted. The expression patterns of the obtained sRNAs were further investigated to detect the differentially expressed (DE) sRNAs between genic male sterile and fertile individuals. A total of 969 DE sRNAs were obtained and only three known miRNA families were included among them. These results suggest that both conserved and species-specific sRNAs contribute to the development of male strobili in <i>C</i>. <i>japonica</i>.</p></div

Pedigree chart for materials used in this study.

<p>Shaded squares represent male sterile individuals expected to be homozygous for <i>ms1</i>.</p

The small RNAs (sRNAs) related to the predicted repeat sequences of <i>Cryptomeria japonica</i>.

<p>The small RNAs (sRNAs) related to the predicted repeat sequences of <i>Cryptomeria japonica</i>.</p

The numbers of DE sRNAs at each sampling date.

<p>The numbers of DE sRNAs at each sampling date.</p

Length distribution of <i>Cryptomeria japonica</i> small RNAs (sRNAs).

<p>Frequency of each size class is shown as a percentage of the total number of retained_sRNAs (black bars) and all unique sequences (white bars).</p