Context: Indian scientists have developed the first-ever low-pungent mustard that is pest and disease-resistant. It is based on CRISPR/Cas9 gene editing, and the crop is non-GM and transgene-free.
Brief background on Oilseed production in India:
- Oilseeds yield oil for cooking, and also their meal – the residual cake after extraction of oil from the seeds – is a protein-rich ingredient used in livestock, poultry and aqua feed.
- Mustard and Soyabean are India’s most widely-cultivated oilseed crops, planted annually on 9 million and 12.5 million hectares area respectively.
- As per the US Department of Agriculture’s data for the marketing year ending September 2023, the share of rapeseed-mustard in India’s production of vegetable oils has been estimated at 42.6% (more than soyabean’s 19.2%) and in that of the meal is 30.3% (next to soyabean’s 38.9%).
- Mustard’s higher average oil extractable content (38% versus 18% for Soyabean) makes it the most significant domestically-grown oilseed crop, while a source of both fats for humans and protein for animals.
- India is a huge importer of edible oils. These imports were valued at $20.84 billion (Rs 167,270 crore) during the fiscal year ended March 2023, while meeting over 60% of the country’s consumption requirement.
Hence, given the massive foreign exchange outgo on account of imports, there is a dire need to boost domestic oilseeds production through focused breeding for improving crop yields, pest and disease resistance, and product quality.
Issue with Mustard:
- Mustard seeds have high levels of glucosinolates, a group of sulphur and nitrogen-containing compoundscontributing to the characteristic pungency of their oil and meal.
- This characteristic pungency limits the oil’s acceptability among consumers due to its strong flavour and odour.
- Also, Rapeseed meal is unpalatable to poultry and pigs, while having to be mixed with fodder grass and water for giving to cattle and buffaloes. High glucosinolates are also known to cause goitre and internal organ abnormalities in livestock.
Breeding GE Canola-quality Mustard (GE low-seed high-leaf glucosinolate Mustard):
1. Brief Background:
- For the past two decades, the scientists at Delhi University’s Centre for Genetic Manipulation of Crop Plants (CGMCP) and the Indian Council of Agricultural Research have been putting efforts into the breeding of rapeseed-mustard lines of so-called Canola quality.
- The dry seeds from the normal mustard (Brassica juncea) cultivated in India contain 120-130 parts per million (ppm or mg/kg) of glucosinolates.
- Whereas the canola quality mustard has below 30 ppm levels of glucosinolates, hence, they are less pungent.
- By lowering the glucosinolate content to the same dry seed weight concentration, the scientists have bred mustard lines whose oil and meal match the standard of canola-quality rapeseed (Brassica napus) in terms of pungency.
- Challenges: However, large-scale cultivation of these canola-quality low-glucosinolate mustard lines has not taken place, because of their vulnerability to pests and diseases. This is because the glucosinolates act as the key arsenal of the Brassicaceae family crops (mustard and canola to cabbage, cauliflower and broccoli) against invading pests, pathogens and termites.
Hence, while the lowering of glucosinolate levels in the seed is desirable for oil and meal, the reduction of its levels in the plant weakens its defence.
2. The Gene Editing breakthrough:
- The scientists have “edited” 10 out of the 12 GTR genes in ‘Varuna’, the high-yielding Indian mustard variety, using CRISPR/Cas9 gene-editing tool.
- CRISPR/Cas9 is a gene-editing tool deploying an enzyme, which acts as a “molecular scissors” to cut the DNA at precisely targeted locations of the gene, and then letting the natural DNA repair process take over.
- In CRISPR genome editing, the first step is the identification of the DNA sequence to be edited.
- Once identified, a guide RNA molecule brings Cas9 protein to the specific spot in the DNA.
- Cas9 protein acts like pair of molecular scissors and cuts the DNA at that spot, and then DNA can be added/removed or edited to correct any mistakes in its sequence.
- CRISPR/Cas9 is a gene-editing tool deploying an enzyme, which acts as a “molecular scissors” to cut the DNA at precisely targeted locations of the gene, and then letting the natural DNA repair process take over.
- In mustard plants, Glucosinolates are synthesised in the leaves and pod walls of the plants. Their translocation and accumulation in the seeds happens through the action of glucosinolate transporter or GTR genes. There are 12 such genes under two distinct classes of GTR1 and GTR2 with six copies each.
- Hence, CRISPR/Cas9 editing was used to create mutations and change the nucleotide sequence of the DNA in the 10 target GTR genes. By doing this, their encoded proteins, responsible for the transport of the glucosinolates to the seeds, were rendered non-functional.
- Results:
- The seeds of the resultant targeted genes-edited Varuna mustard variety had glucosinolate content well below the 30 ppm canola-quality threshold.
- At the same time, the other plant parts, especially the leaves and pod walls enclosing the seeds, had significantly higher glucosinolate accumulation, and hence, the defence response was at par with or better than that of the wild-type mustard.
Genetically-modified crops vs Genetically-engineered crops:
- The low-seed high-leaf glucosinolate mustard lines developed are genome edited or GE, as opposed to GM or transgenic plants.
- The new GTR genes-edited mustard lines are transgene-free or non-genetically modified (GM).
- They contain no foreign genes like those of the Bacillus thuringiensis bacteria in cotton or Bar-Barnase-Barstar (isolated from other soil bacteria) in the GM hybrid mustard (DMH-11).
GM vs GE:
1. Process:
- Genetically-Modified Crops (GMOs): Often abbreviated as GMOs (genetically modified organisms), the genetic material of such crops has been altered through a process of genetic engineering. This modification can involve the introduction of genes from other organisms to confer specific traits, such as resistance to pests, tolerance to herbicides, improved nutritional content, or longer shelf life. GMOs can be created using various techniques, including transgenesis (the insertion of genes from one species into another) and cisgenesis (transfer of genes within the same species or a closely related one). E.g., DMH-11
- Genetically-Engineered Crops: Genetically-engineered crops encompass a broader category that includes both GMOs and those created through genome editing. The Genome editing techniques, such as CRISPR-Cas9, allow for more precise modifications to the DNA of an organism. This technique can involve adding, deleting, or modifying specific DNA sequences within an organism’s genome.
2. Clearance:
- GM crops are currently subjected to stringent “environmental release” regulations in India, not just for commercial cultivation but even for field trials and seed production. Such release is contingent upon clearance from a special Genetic Engineering Appraisal Committee (GEAC) under the Ministry of Environment, Forest and Climate Change (MoEFCC). The GEAC’s green signal is itself not binding on the Union Government, which gives the final nod.
- On March 30, 2022, an office memorandum from the MoEFCC exempted GE plants “free of exogenous introduced DNA” from the requirement of GEAC approval for open field trials leading to commercial release. Such clearance is now necessary only at the level of an Institutional Bio-safety Committee (IBSC), comprising scientists from the institutions engaged in the GE crop development and from the DBT.
Hence, the scientists are hopeful of conducting open field trials of the new GE mustard line in the coming rabi planting season (October-November).
The new GE low-seed high-leaf glucosinolate lines and GM hybrid mustard (like DMH-11) are major plant breeding advancements – from Indian scientists – that can go some way towards bringing down the dependence on imported vegetable oils.
