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Transient expression from microprojectile-mediated DNA transfer in pinus taeda.

A M Stomp1, A Weissinger, R R Sederoff

  • 1Department of Forestry, North Carolina State University, 27695, Raleigh, NC, USA.

Plant Cell Reports
|November 14, 2013
PubMed
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Microprojectile bombardment successfully introduced plasmid DNA into loblolly pine cells, showing transient beta-glucuronidase (GUS) gene expression. While gene expression was detected long-term, transgenic loblolly pine (Pinus taeda L.) plants were not recovered.

Area of Science:

  • Plant Biotechnology
  • Molecular Biology
  • Forest Genetics

Background:

  • Efficient genetic transformation of coniferous species like loblolly pine (Pinus taeda L.) is crucial for forest improvement.
  • Microprojectile bombardment is a physical method for delivering foreign DNA into plant cells.

Purpose of the Study:

  • To demonstrate the feasibility of gene transfer into loblolly pine cotyledon cells using microprojectile bombardment.
  • To assess transient gene expression and cellular effects following DNA delivery.

Main Methods:

  • Loblolly pine cotyledon cells were bombarded with plasmid DNA encoding the beta-glucuronidase (GUS) reporter gene.
  • GUS activity was detected using histochemical staining at various time points post-bombardment.
  • Cellular viability and damage were assessed in GUS-positive cells.

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Main Results:

  • Transient GUS expression was observed in localized centers (blue spots) within 24 hours.
  • Expression persisted in meristematic tissue up to 62 days, though at declining levels.
  • Expression events primarily originated from single-cell transformations, with complex staining patterns in adjacent cells.
  • Significant cellular damage was observed in some GUS-positive cells, ranging from minor to lethal.

Conclusions:

  • Microprojectile bombardment is an effective tool for assaying transient gene expression in loblolly pine.
  • The method shows potential for developing transgenic pine plants, despite challenges in recovering whole transgenic shoots.
  • Further optimization is needed to improve transformation efficiency and minimize cellular damage for successful plant regeneration.