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Over-Expression of the Cell-Cycle Gene LaCDKB1;2 Promotes Cell Proliferation and the Formation of Normal Cotyledonary Embryos during Larix kaempferi Somatic Embryogenesis.

ABSTRACT: Somatic embryogenesis is an effective tool for the production of forest tree seedlings with desirable characteristics; however, the low initiation frequency and productivity of high-quality mature somatic embryos are still limiting factors for Larix kaempferi (Japanese larch). Here, we analyzed the expression pattern of L. kaempferi cyclin-dependent kinase B 1;2 (LaCDKB1;2) during somatic embryogenesis in L. kaempferi and its relationship with the cell proliferation rate. We also analyzed the effect of LaCDKB1;2 over-expression on somatic embryo quality. The results revealed a positive correlation between LaCDKB1;2 expression and the cell proliferation rate during the proliferation stage. After LaCDKB1;2 over-expression, the proliferation rate of cultures increased, and the number of somatic embryos in transgenic cultures was 2.69 times that in non-transformed cultures. Notably, the number of normal cotyledonary embryos in transgenic cultures was 3 times that in non-transformed cultures, indicating that LaCDKB1;2 not only increases the proliferation of cultures and the number of somatic embryos but also improves the quality of somatic embryos. These results provide insight into the regulatory mechanisms of somatic embryogenesis as well as new Larix breeding material.

SUBMITTER: Kang Y

PROVIDER: S-EPMC8468589 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Hybrids of larch (Larix kaempferi × Larix olgensis) are important afforestation species in northeastern China. They are routinely propagated via rooted stem cuttings. Despite the importance of rooting, little is known about the regulation of adventitious root development in larch hybrids. 454 GS FLX Titanium technology represents a new method for characterizing the transcriptomes of non-model species. This method can be used to identify differentially expressed genes, and then two-dimensional difference gel electrophoresis (2D-DIGE) and matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF/TOF MS) analyses can be used to analyze their corresponding proteins. In this study, we analyzed semi-lignified cuttings of two clones of L. kaempferi × L. olgensis with different rooting capacities to study the molecular basis of adventitious root development.We analyzed two clones; clone 25-5, with strong rooting capacity, and clone 23-12, with weak rooting capacity. We constructed four cDNA libraries from 25-5 and 23-12 at two development stages. Sequencing was conducted using the 454 pyrosequencing platform. A total of 957832 raw reads was produced; 95.07% were high-quality reads, and were assembled into 45137 contigs and 61647 singletons. The functions of the unigenes, as indicated by their Gene Ontology annotation, included diverse roles in the molecular functions, biological processes, and cellular component categories. We analyzed 75 protein spots (-fold change ? 2, P ? 0.05) by 2D-DIGE, and identified the differentially expressed proteins using MALDI-TOF/TOF MS. A joint analysis of transcriptome and proteome showed genes related to two pathways, polyamine synthesis and stress response, might play an important role on adventitious root development.These results provide fundamental and important information for research on the molecular mechanism of adventitious root development. We also demonstrated for the first time the combined use of two important technologies as a powerful approach to advance research on non-model, but otherwise important, larch species.

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Over-Expression of the Cell-Cycle Gene LaCDKB1;2 Promotes Cell Proliferation and the Formation of Normal Cotyledonary Embryos during Larix kaempferi Somatic Embryogenesis.

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