Samnakay, Parwez and Davies, Bill (2015) An investigation into the chloroplast transformation of wheat, and the use of a cyanobacterial CCM gene for improving photosynthesis in a C3 plant. PhD thesis, Lancaster University.
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Abstract
Wheat is a major component of the UK diet, and provides approximately 20% of global caloric intake. Wheat is grown on more land area than any other crop, and the continued supply of wheat is essential for global food security. Biotechnology is likely to play an important role in the sustainable increase of wheat yields, and the genetic manipulation of chloroplasts for photosynthetic improvement has many potential advantages over transformation of the nuclear genome. The genetic modification of the chloroplast genome via transformation was first demonstrated in the late 1980’s, and since then, chloroplast transformation of many Dicotyledonous (dicot) plant species such as Nicotiana tabaccum has been routinely performed. In comparison, the transformation of chloroplasts in Monocotyledons (monocot) plant species, which includes all cereal crops, has made far less progress. To date, there has been no reproducible homoplasmic plastid transformation event in the monocots. This study identifies a number of bottlenecks responsible for the prevention of chloroplast transformation in wheat. One such bottleneck is the lack of a suitable explant for plastid transformation, as traditional nuclear transformation targets are absent of metabolically active plastids. This study has developed a robust regeneration protocol for a previously undescribed tissue, termed the primary inflorescence leaf sheath (piLS), which is rich in active chloroplasts. Functional wheat specific chloroplast transformation vectors have been generated, and bombardment studies have been conducted with these on piLS and a second tissue, the immature embryosderived callus. Immature embryo callus (IEC) does not contain active plastids, however contains pro-plastids and is highly embryogenic. To uncover novel ways of increasing photosynthesis in C3 plants, a number of transplastomic tobacco lines expressing the Synechococcus elongatus PCC 7942 ictB gene were generated. Previous studies suggest that ictB may be an inorganic carbon transporter. In a number of transplastomic lines produced in this study, the intercellular carbon concentration (Ci) is significantly increased. This increased Ci did not result in an increased photosynthetic rate, however did cause a number of phenotypic differences, such as smaller plants, wider leaves, and earlier seed pod formation. The results, with regards to chloroplast transformation, and its implications in improving photosynthesis within C3 plants, are discussed in this thesis.