High-throughput sequencing of small RNAs from seedling of Larix leptolepis
Ontology highlight
ABSTRACT:
ORGANISM(S): Larix kaempferi
PROVIDER: GSE34805 | GEO | 2012/12/28
SECONDARY ACCESSION(S): PRJNA151137
REPOSITORIES: GEO
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Project description:MicroRNAs (miRNAs) are emerging as essential, albeit poorly characterized, regulators of biological processes. The miRNA in gymnosperms is under-identified, which limits the progress of miRNA in gymnoperms. Using the high-throughput sequencing, a total of 87 conserved miRNAs were identified from Larix leptolepis. Eighteen novel miRNAs were discovered in our library, most of which were Larix-specific miRNAs. Identification of small RNA in Larix seedling
2012-12-28 | E-GEOD-34805 | biostudies-arrayexpress
Project description:MicroRNAs (miRNAs) are emerging as essential regulators of biological processes. Somatic embryogenesis is one of the most important techniques for gymnosperm breeding programs, but there is little understanding of its underlying mechanism. To investigate the roles of miRNAs during somatic embryogenesis in larch, we constructed a small RNA library from somatic embryos. High-throughput sequencing of the library identified 83 conserved miRNAs from 35 families, 16 novel miRNAs, and 14 plausible miRNA candidates, with a high proportion specific to larch or gymnosperms. qRT-PCR analysis demonstrated that both the conserved and novel or candidate miRNAs were expressed in larch. Several miRNA precursor sequences were obtained via RACE. We predicted 110 target genes using bioinformatics, and validated nine of them by 5M-bM-^@M-^Y RACE. Eleven conserved miRNA families including 17 miRNAs with critical functions in plant development and six target mRNAs were detected by qRT-PCR in the larch SE. Stage-specific expression of miRNAs and their targets indicate their possible modulation on SE of larch: miR171a/b might exert function on PEMs, while miR171c acts in the induction process of larch SE; miR397 and miR398 mainly involved in modulation of PEM propagation and transition to single embryo; miR162 and miR168 exert their regulatory function during total SE process, especially during stage 5 to stage 8; miR156, miR159, miR160, miR166, miR167, and miR390 might play regulatory roles during cytoledonary embryo development. These findings indicate that larch and possibly other gymnosperms have complex mechanisms of gene regulation involving specific and common miRNAs operating post-transcriptionally during embryogenesis. Examination of small RNA expression profilings in Larix somatic embryos over seven developmental stages: 1) after 2 and 15 days on sub-culture; 2) 2, 6, and 11 days post-differentiation culture; and 3) somatic embryos at days 33 and 39.
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Project description:To investigate the roles of sRNAs in keeping embryo dormancy or germination in Larix leptolepis, we deciphered the endogenous "sRNAome" in dormant and germinated embryos. High-throughput sequencing of the sRNA libraries showed that dormant embryos exhibited a length bias towards 24-nt, while germinated embryos showed a bias towards a 21-nt and/or 22-nt length. Both of proportions for miRNAs to the non-redundant and redundant sRNAs were higher in germinated embryos than those in dormant embryos, while the ratio of unknown sRNAs was higher in dormant embryos than in germinated embryos. The proportion of 21-nt and 22-nt sRNAs increased in germinated embryos, which might attribute to the higher expression level of miRNAs. We identified a total of 160 conserved miRNAs from 39 families, 16 novel miRNAs, and 14 plausible miRNA candidates, of which novel and non-conserved known miRNAs might be the main contributors. These findings indicate that larch and possibly other gymnosperms have complex mechanisms of gene regulation involving sRNAs and miRNAs operating transcriptionally and post-transcriptionally during embryo dormancy and germination.
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Project description:Small non-coding RNAs (sncRNAs) are emerging as key regulators of embryogenesis. To investigate the roles of sRNAs in regulating synchronism of somatic embryogenesis in Larix leptolepis, we deciphered the endogenous "sRNAome" in synchronous and desynchronous embryos. The 24-nt class sRNAs were overrepresented in both synchronous embryos and desynchronous embryos, accounting for 85.29% and 44.79%. A total of 29 miRNAs were upregulated in synchronous embryos, whereas 59 miRNAs were upregulated in desynchronous embryos. We describe the emerging theme for sncRNAs function: inhibiting the precocious expression, thus regulating the synchronism of somatic embryogenesis. These findings indicate that larch and possibly other gymnosperms have complex mechanisms of gene regulation involving sRNAs and miRNAs operating transcriptionally and post-transcriptionally during the regulation of synchronism.
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Project description:To investigate the roles of sRNAs in keeping embryo dormancy or germination in Larix leptolepis, we deciphered the endogenous "sRNAome" in dormant and germinated embryos. High-throughput sequencing of the sRNA libraries showed that dormant embryos exhibited a length bias towards 24-nt, while germinated embryos showed a bias towards a 21-nt and/or 22-nt length. Both of proportions for miRNAs to the non-redundant and redundant sRNAs were higher in germinated embryos than those in dormant embryos, while the ratio of unknown sRNAs was higher in dormant embryos than in germinated embryos. The proportion of 21-nt and 22-nt sRNAs increased in germinated embryos, which might attribute to the higher expression level of miRNAs. We identified a total of 160 conserved miRNAs from 39 families, 16 novel miRNAs, and 14 plausible miRNA candidates, of which novel and non-conserved known miRNAs might be the main contributors. These findings indicate that larch and possibly other gymnosperms have complex mechanisms of gene regulation involving sRNAs and miRNAs operating transcriptionally and post-transcriptionally during embryo dormancy and germination. One embryogenic cell line of Japanese larch (Larix leptolepis), designated as D878, with a high embryo maturation capacity was used in this study. Embryogenic callus were induced from immature embryos of larch on induction medium, followed by sub-culture, and culture on ABA-containing mature medium in a dark environment at 25 2 C. After cultured 45 days in mature medium, embryogenic calli developed into mature somatic embryos. In our study, the samples were harvested at day 57, one sample was collected after mature embryos continued to stay for 12 days on ABA-containing medium, and the other one was harvested after cultured for 12 days on ABA-removing medium. All samples were snap-frozen in liquid nitrogen, and stored in liquid nitrogen until RNA extraction.
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Project description:Small non-coding RNAs (sncRNAs) are emerging as key regulators of embryogenesis. To investigate the roles of sRNAs in regulating synchronism of somatic embryogenesis in Larix leptolepis, we deciphered the endogenous "sRNAome" in synchronous and desynchronous embryos. The 24-nt class sRNAs were overrepresented in both synchronous embryos and desynchronous embryos, accounting for 85.29% and 44.79%. A total of 29 miRNAs were upregulated in synchronous embryos, whereas 59 miRNAs were upregulated in desynchronous embryos. We describe the emerging theme for sncRNAs function: inhibiting the precocious expression, thus regulating the synchronism of somatic embryogenesis. These findings indicate that larch and possibly other gymnosperms have complex mechanisms of gene regulation involving sRNAs and miRNAs operating transcriptionally and post-transcriptionally during the regulation of synchronism. One embryogenic cell line of Japanese larch (Larix leptolepis), designated as D878, with a high embryo maturation capacity was used in this study. Embryogenic callus was induced from immature embryos of Japanese larch on induction medium followed by sub-culture. Calli at the proembryogenic mass III stage were cultured on maturation medium in a dark environment at 25 M-BM-1 2M-BM-0C. Samples were cultured on ABA-plus or ABA-minus maturation medium for 45 days. All samples were snap-frozen in liquid nitrogen, and stored in liquid nitrogen until RNA extraction.
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