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Comparison of gene expression markers during zygotic and somatic embryogenesis in pine

Abstract

The relative expression of developmentally regulated genes was analyzed during zygotic embryo development in Pinus taeda and somatic embryo development/maturation in P. taeda and Pinus oocarpa. The following four embryo samples were studied: zygotic pro-embryos and somatic embryogenic suspensor masses; round/globular, early cotyledonary, and late cotyledonary. The relative transcript levels of six genes of interest: legumin-/vicilin-like, group 4 late embryogenesis abundant, homeodomain-leucine zipper I, 26S proteasome regulatory subunit S2, and clavata-like, associated with different aspects of embryo development, were analyzed by real-time PCR. In both pine species, the relative transcript levels for legumin-/vicilin-like storage proteins and the late embryogenesis abundant protein accumulated gradually through somatic embryo maturation, in contrast to zygotic embryos, where transcripts increased significantly to their highest levels at the late cotyledonary stage. The homeodomain-leucine zipper I relative transcript accumulation pattern differed between somatic and zygotic embryos. This difference was attributed to differences in cell-type compositions between the embryogenic suspensor masses and pro-embryos. Additionally, in P. oocarpa, the transcript levels of homeodomain-leucine zipper I remained high after the transfer of somatic embryos to maturation conditions, but declined during maturation in P. taeda. The highest 26S proteasome regulatory subunit S2 relative transcript levels in P. taeda were in round/globular somatic and zygotic embryos and in embryogenic suspensor masses and round/globular P. oocarpa somatic embryos. Finally, the relative transcripts levels for the clavata-like gene were more predominant in round/globular, early cotyledonary, and embryogenic suspensor masses in P. taeda and P. oocarpa, respectively. Somatic embryos exhibited relative gene expression patterns similar to their zygotic counterparts, although some differences were noted between zygotic and somatic embryos, as well as between the different pine somatic embryo systems.

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Acknowledgments

This work represents a portion of A. Lara-Chavez’s doctoral dissertation. We would like to thank Dr. Scott Merkle for providing P. taeda cell cultures, Kyle Peer for collecting the immature P. taeda cones, Dr. Marty Fernandes for providing developing P. oocarpa cones, Anne Kathryn Dalton for help in embryo dissection and collection, Dr. Supriya Ratnaparkhe for providing the photo of the P. taeda pro-embryo stage in Fig. 1 and Guozhu Tang for support with RNA extraction and cDNA synthesis. Many thanks also to Drs. Scott Merkle, Thomas Fox and Jose Alexander Elvir for their comments during the course of this study. This work was funded through Special Grants (2003-38891-02112, 2008-38891-19353 and 2009-38891-20092) and HATCH funds (Project No. VA-135816) from the United States Department of Agriculture, and operating funds from the Commonwealth of Virginia to the Institute for Advanced Learning and Research.

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Correspondence to Barry S. Flinn.

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Ulrika Egertsdotter and Barry S. Flinn contributed equally to project development, support, and supervision.

Editor: Thomas Clemente

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Lara-Chavez, A., Egertsdotter, U. & Flinn, B.S. Comparison of gene expression markers during zygotic and somatic embryogenesis in pine. In Vitro Cell.Dev.Biol.-Plant 48, 341–354 (2012). https://doi.org/10.1007/s11627-012-9440-5

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  • DOI: https://doi.org/10.1007/s11627-012-9440-5

Keywords

  • Pinus
  • Storage protein
  • LEA
  • Clavata-like
  • RPN1
  • HD-Zip I