Development of imidazolinone herbicide tolerant borage (Borago officinalis L.).

Borage (Borago officinalis) is an annual herb that produces a high level of gamma-linolenic acid (GLA) in its seed oil. Due to the recognized health benefits of GLA, borage is now commercially cultivated worldwide. However, an herbicide-tolerant variety for effective weed management has not yet been developed. Here we report the generation and characterization of ethyl methanesulfonate (EMS) induced borage mutant lines tolerant to the herbicide imidazolinone. An EMS-mutagenized borage population was generated by using a series of concentrations of EMS to treat mature borage seeds. Screening of the M2 and M3 borage plants using an herbicide treatment resulted in the identification of two imidazolinone-tolerant lines. Sequence analysis of two acetohydroxyacid synthase (AHAS) genes, AHAS1 and AHAS2, from the mutant (tolerant) and wild type (susceptible) borage plants showed that single nucleotide substitutions which resulted in amino acid changes occurred in AHAS1 and AHAS2, respectively in the two tolerant lines. A KASP marker was then developed to differentiate the homozygous susceptible, homozygous tolerant and heterozygous borage plants. An in vitro assay showed that homozygous tolerant borage carrying the AHAS1 mutation retained significantly higher AHAS activity than susceptible borage across different imazamox concentrations. A herbicide dose response test indicated that the line with the AHAS1 mutation could tolerate four times the normally used field concentration of "Solo" herbicide.

Delta-6 desaturase from borage converts linoleic acid to gamma-linolenic acid in HEK293 cells.

Gamma-linolenic acid (GLA, 18:3 n6) is an essential polyunsaturated fatty acid of the omega-6 family and is found to be effective in prevention and/or treatment of various health problems. In this study, we evaluated the possibility of increasing γ-linolenic acid contents in mammalian cells using the delta-6 gene from Borago officinalis. The borage Δ6-desaturase gene (sDelta-6) was codon-optimized and introduced into HEK293 cells by lipofectin transfection. Co-expression of GFP with sDelta-6 and RT-PCR analysis indicated that sDelta-6 could be expressed in mammalian cells. Subsequently, the heterologous expression of borage Δ6-desaturase was evaluated by fatty acid analysis. Total cellular lipid analysis of transformed cells fed with linoleic acid (LA 18:2 n6) as a substrate showed that the expression of sDelta-6 resulted in an 228-483% (p<0.05) increase of GLA when compared with that in the control cells. The highest conversion efficiency of LA into GLA in sDelta-6(+) cells was 6.9 times higher than that in the control group (11.59% vs. 1.69%; p<0.05). Our present work demonstrated that the sDelta-6 gene from borage could be functionally expressed in mammalian cells, and could convert LA into GLA. Furthermore, this study may pave the way to generate transgenic livestock that can synthesise GLA.

Modification of fatty acid composition in tomato (Lycopersicon esculentum) by expression of a borage delta6-desaturase.

The improvement of nutritional quality is one potential application for the genetic modification of plants. One possible target for such manipulation is the modification of fatty acid metabolism. In this work, expression of a borage delta6-desaturase cDNA in tomato (Lycopersicon esculentum L.) has been shown to produce gamma-linolenic acid (GLA; 18:83 delta6,9,12) and octadecatetraenoic acid (OTA; 18:4 delta6,9,12,15) in transgenic leaf and fruit tissue. This genetic modification has also, unexpectedly, resulted in a reduction in the percentage of linoleic acid (LA 18:2 delta9,12) and a concomitant increase in the percentage of alpha-linolenic acid (ALA; 18:3 delta9,12,15) in fruit tissue. These changes in fatty acid composition are thought to be beneficial for human health.