Dataset Information

Simple Culture Methods and Treatment to Study Hormonal Regulation of Ovule Development.

ABSTRACT: Ovule development is one of the most important processes in the reproductive development of higher plants and is a determinant of seed quality and quantity. Phytohormones play key roles in this process since loss-of-function mutants in hormone signaling show defective ovule phenotypes and reduced fertility. However, it is difficult to distinguish the direct effects of hormones on ovule development because it is parts of reproductive development and the defective phenotypes would be the indirect effects following the defective vegetative development. The treatment of hormones is a direct method to investigate the hormonal regulation of ovule development, but ovule is embedded inside several layers of floral organs, and traditional methods for hormone (or inhibitor) treatments have various limitations. We have developed simple methods to apply treatments to the flowers in a living plant, where an inflorescence apex is immersed into a solution in an inverted tube. We have also developed a specific system to culture and treat excised flowers/pistils. These procedures will be useful for research on the hormonal regulation of ovule development. We provide examples of how treatments with brassinosteroids (BR) and BR biosynthesis inhibitor. We cultured and treated plant materials using our newly developed methods, and observed the morphology of wild type ovules and fluorescence signals in a marker line to monitor the progress of ovule development. The results demonstrate BR promotes ovule development and our new methods are efficient and repeatable.

SUBMITTER: Li BF

PROVIDER: S-EPMC6015908 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Simple Culture Methods and Treatment to Study Hormonal Regulation of Ovule Development.

Li Bu-Fan BF Yu Shi-Xia SX Hu Li-Qin LQ Zhang Yan-Jie YJ Zhai Ning N Xu Lin L Lin Wen-Hui WH

Frontiers in plant science 20180618

Ovule development is one of the most important processes in the reproductive development of higher plants and is a determinant of seed quality and quantity. Phytohormones play key roles in this process since loss-of-function mutants in hormone signaling show defective ovule phenotypes and reduced fertility. However, it is difficult to distinguish the direct effects of hormones on ovule development because it is parts of reproductive development and the defective phenotypes would be the indirect ...[more]

PMID: 29967629

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Project description:BACKGROUND: The involvement of small RNAs in cotton fiber development is under explored. The objective of this work was to directly clone, annotate, and analyze small RNAs of developing ovules to reveal the candidate small interfering RNA/microRNAs involved in cotton ovule and fiber development. RESULTS: We cloned small RNA sequences from 0-10 days post anthesis (DPA) developing cotton ovules. A total of 6691 individual colonies were sequenced from 11 ovule small RNA libraries that yielded 2482 candidate small RNAs with a total of 583 unique sequence signatures. The majority (362, 62.1%) of these 583 sequences were 24 nt long with an additional 145 sequences (24.9%) in the 21 nt to 23 nt size range. Among all small RNA sequence signatures only three mirBase-confirmed plant microRNAs (miR172, miR390 and ath-miR853-like) were identified and only two miRNA-containing clones were recovered beyond 4 DPA. Further, among all of the small RNA sequences obtained from the small RNA pools in developing ovules, only 15 groups of sequences were observed in more than one DPA period. Of these, only five were present in more than two DPA periods. Two of these were miR-172 and miR-390 and a third was identified as 5.8S rRNA sequence. Thus, the vast majority of sequence signatures were expressed in only one DPA period and this included nearly all of the 24 nt sequences. Finally, we observed a distinct DPA-specific expression pattern among our clones based upon sequence abundance. Sequences occurring only once were far more likely to be seen in the 0 to 2 DPA periods while those occurring five or more times were the majority in later periods. CONCLUSION: This initial survey of small RNA sequences present in developing ovules in cotton indicates that fiber development is under complex small RNA regulation. Taken together, the results of this initial small RNA screen of developing cotton ovules is most consistent with a model, proposed by Baulcombe, that there are networks of small RNAs that are induced in a cascade fashion by the action of miRNAs and that the nature of these cascades can change from tissue to tissue and developmental stage to developmental stage.

Project description:BackgroundArabidopsis ovules comprise four morphologically distinct parts: the nucellus, which contains the embryo sac, two integuments that become the seed coat, and the funiculus that anchors the ovule within the carpel. Analysis of developmental mutants has shown that ovule morphogenesis relies on tightly regulated genetic interactions that can serve as a model for developmental regulation. Redundancy, pleiotropic effects and subtle phenotypes may preclude identification of mutants affecting some processes in screens for phenotypic changes. Expression-based gene discovery can be used access such obscured genes.ResultsAffymetrix microarrays were used for expression-based gene discovery to identify sets of genes expressed in either or both integuments. The genes were identified by comparison of pistil mRNA from wild type with mRNA from two mutants; inner no outer (ino, which lacks the outer integument), and aintegumenta (ant, which lacks both integuments). Pools of pistils representing early and late stages of ovule development were evaluated and data from the three genotypes were used to designate genes that were predominantly expressed in the integuments using pair-wise and cluster analyses. Approximately two hundred genes were found to have a high probability of preferential expression in these structures, and the predictive nature of the expression classes was confirmed with reverse transcriptase polymerase chain reaction and in situ hybridization.ConclusionThe results showed that it was possible to use a mutant, ant, with broad effects on plant phenotype to identify genes expressed specifically in ovules, when coupled with predictions from known gene expression patterns, or in combination with a more specific mutant, ino. Robust microarray averaging (RMA) analysis of array data provided the most reliable comparisons, especially for weakly expressed genes. The studies yielded an over-abundance of transcriptional regulators in the identified genes, and these form a set of candidate genes for evaluation of roles in ovule development using reverse genetics.

Dataset Information

Simple Culture Methods and Treatment to Study Hormonal Regulation of Ovule Development.

Publications

Simple Culture Methods and Treatment to Study Hormonal Regulation of Ovule Development.

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