1. STEPS IN DEVELOPMENT OF TRANSGENIC CROPS

2. REGULATORY APPROVAL SYSTEM OF BIOTECH CROPS

Presentation by: Guru P N 2015-16

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What is a transgenic?

Transgene – the genetically engineered gene added to a species

Example: – modified EPSP synthase gene (encodes a protein that functions even when plant is treated with Roundup)

Transgenic – an organism containing a transgene introduced by technological (not breeding) methods

Example:– Roundup Ready Crops

Concept Based on the Term Transgene

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We can develop organisms that express a “novel” trait not normally found in the species

Why are transgenics important?

Extended shelf-life tomato (Flavr-Savr)

Herbicide resistant soybean (Roundup Ready)

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Insect resistant transgenics

Source: USDA

Insect resistant cotton – Bt toxin kills the cotton boll worm• transgene = Bt protein

Insect resistant corn – Bt toxin kills the European corn borer• transgene = Bt protein

Normal Transgenic

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Plant Transformation Methods

Physical Chemical Biological

MicroinjectionPressureBiolistics - gene gun/particle bombardmentElectroporationMicroinjection Silica/carbon fibersLazer mediated SAT

PEGDEAE-dextranCalcium phosphateArtificial lipidsProteinsDendrimers

A. TumefaciensA. Rhizogenes

Virus-mediated

In planta

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Important Plant Improvement Methods

• Breeding Crossing two individuals from the same species; produces a new, improved variety; not a biotechnology procedure

• Transformation Adding a gene from another species; the essential biotechnology procedure to produce transgenics

Source: USDA

Source: USDA

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Introducing the Geneor

Developing Transgenics

Steps

1. Create transformation cassette

2. Introduce and select for transformants

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Transformation Cassettes

Contains

1. Gene of interest• The coding region and its controlling elements

2. Selectable marker• Distinguishes transformed/untransformed plants

3. Insertion sequences• Aids Agrobacterium insertion

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Transformation Steps

Prepare tissue for transformation

Introduce DNA

Culture plant tissue• Develop shoots• Root the shoots

Field test the plants

• Leaf, germinating seed, immature embryos• Tissue must be capable of developing into normal plants

• Agrobacterium or gene gun

• Multiple sites, multiple years

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• Transformation cassettes are developed in the lab

• They are then introduced into a plant

• Two major delivery methods

Delivering the Geneto the Plant

• Agrobacterium

• Gene GunTissue culturerequired to generatetransgenic plants

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The Lab Steps

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Production of transgenic plants

Isolate and clone gene of interest

Add DNA segments to initiate or enhance gene expression

Add selectable markers

Introduce gene construct into plant cells (transformation)

Select transformed cells or tissues

Regenerate whole plants15

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BIOTECHNOLOGY REGULATORY AUTHORITY OF INDIA

ORGANOGRAM OF BRAI

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Regulation of GM Foods in India

Under Environment Protection Act (1986), MoEf rules formulated in 1989 which created six competent authorities namely RDAC, RCGM, GEAC, IBSC, SBCC, DLC;

MEC for field trials under RCGM and GEAC Recombinant DNA Safety Guidelines 1990, modified 1994 DBT Revised Guidelines for research on Transgenic Plants –

1998, also includes guidelines for Toxicity & Allergenecity evaluation of transgenic seeds, plants & plant parts

Safety assessment of GM Food Crops –2007, modified 2009 Guidelines for the safety assessment of foods derived from

GE plants -2008 Guidelines & SOPs for Confined Field Trials of Regulated

Genetically Engineered Plants –2008

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Indian Regulatory System

Aims to ensure that GM crops pose no risk to food safety, environmental safety and agriculture productivity To demonstrate potential benefits over the conventional variety/hybrid in terms of economic benefit to the farmer and /or the environment To generate quantitative biological, ecological and agronomic supportive data Rules and guidelines formulated to achieve above objectives

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AGENDA

Gene Stacking Crop Rotation Bt as part of IPM Agricultural practices to integrate GM crops SiRNA approach Minor Pests Marker-Free S stage-Specific Promoter Improved Nutrition & Abiotic Stress

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GOVERNMENT RULES FOR GMOs

The regulatory framework for transgenic crops in India consists of the following rules and guidelines.

Rules and policies Rules 1989 under Environment Protection Act (1986)

Seed Policy 2002 Guidelines

Recombinant DNA guidelines, 1990

Guidelines for research in transgenic crops, 1998

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Rules, 1989

There are six competent authorities as per the rules:

Recombinant DNA Advisory Committee (RDAC)

Review Committee on Genetic Manipulation (RCGM)

Genetic Engineering Approval Committee (GEAC)

Institutional Biosafety Committees (IBSC)

State Biosafety Coordination Committees (SBCC)

District Level Committees (DLC).

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Out of these, the three agencies that are involved in approval of new transgenic crops are:

IBSC set-up at each institution for monitoring institute level research in genetically modified organ- isms.

RCGM functioning in the DBT to monitor ongoing research activities in GMOs and small scale field trials.

GEAC functioning in the MoEF to authorize large-scale trials and environmental release of GMOs.

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REGULATORY MECHANISMS

3 Tier System

•IBSC

•RCGM-MEC (Monitoring-cum- Evaluation)

•GEAC

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The approvals and prohibitions under Rules 1989 are summarized below:

No person shall import, export, transport, manufacture, process, use or sell any GMOs, substances or cells except with the approval of the GEAC.

Use of pathogenic organisms or GMOs or cells for research purpose shall be allowed under the Notification, 1989 of the EPA, 1986.

Any person operating or using GMOs for scale up or pilot operations shall have to obtain permission from GEAC.

For purpose of education, experiments on GMOs IBSC can look after, as per the guidelines of the Government of India.

Deliberate or unintentional release of GMOs not allowed. Production in which GMOs are generated or used shall not be

commenced except with the approval of GEAC supervises the implementation of rules and guidelines.

GEAC carries out supervision through SBCC, DLC or any authorized person.

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If orders are not complied, SBCC/DLC may take suitable measures at the expenses of the person who is responsible.

In case of immediate interventions to prevent any damage, SBCC and DLC can take suitable measures and the expenses incurred will be recovered from the person responsible.

All approvals shall be for a period of 4 years at fi rst instance renewable for 2 years at a time. GEAC shall have powers to revoke approvals in case of: Any new information on harmful e ects of GMOs. GMOs cause such damage to the ffenvironment as could not be envisaged when approval was given. Non-compliance of any conditions stipulated by GEAC.

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Genetic engineering experiments on plants have been grouped under three categories.

Category I includes routine cloning of defined genes, defined non-coding stretches of DNA and open reading frames in defined genes in E. coli or other bacterial/fungal hosts which are generally considered as safe to human, animals and plants.

Category II experiments include experiments carried out in lab and green house/net house using defined DNA fragments non-pathogenic to human and animals for genetic transformation of plants, both model species and crop species.

Category III includes experiments having high risk where the escape of transgenic traits into the open environment could cause significant alterations in the biosphere, the ecosystem, plants and animals by dispersing new genetic traits the e ects of which cannot be judged precisely. This also ffincludes experiments having risks mentioned above conducted in green houses and open field conditions. 28

OVERVIEW OF MINISTRIES AND DEPARTMENTS INVOLVED IN REGULATION

OF GM FOOD

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1. Ministry of Environment and Forest:

This ministry holds the Secretariat of the Genetic Engineering Approval Committee, the apex body that gives approval for manufacture, sale, import and export of all GMOs and products thereof including foodstu , ingredients in foodstu and additives using genetically ff ffmodifi ed (GM) organisms or cells.

2. Department of Biotechnology:

This department holds the Secretariat of the Review Committee on Genetically Modifi cation that gives approval for research and small scale fi eld trials involving GMOs and products thereof. It also interacts with the Institutional Biosafety Committees (IBSCs) set up in all organizations undertaking activities involves GMOs.

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3. Department of Health in the Ministry of Health and Family Welfare:

Department of Health is responsible for implementation of the PFA Act under which the quality and safety of food is regulated. The Directorate General of Health Services has also been designed as the nodal Ministry with the Codex Alimentarious Commission.

4. The Indian Council of Medical Research (ICMR)

It is the apex body in India for the formulation, coordination and promotion of biomedical research under the Ministry of Health and Family Welfare. ICMR acts as an advisory body for MoHFW on various issues including GM foods.

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5. Ministry of Agriculture:

Ministry of Agriculture is the nodal ministry for agriculture growth in the country. It comprises of three Departments viz. Department of Agriculture and Cooperation, Department of Agricultural Research & Education/ Indian Council of Agricultural Research (ICAR) and Department of Animal Husbandry & Dairying. Th e o cials from ICAR and Ministry of ffiAgriculture have an important role to play in the approval of GM crops as per Seed Policy, 2002.

6. Ministry of Commerce and Industry:

This ministry is responsible for the formulation of the Export and Import (EXIM) Policy in the country. It implements a legislation prescribing a system of quality control and inspection for both export/import.

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7. Ministry of Food Processing Industries:

This ministry is responsible for the formulation of policy for the healthy growth of the food processing industries and provides developmental support to these industries. It encourages research and developmental activities and assists the industries in active participation in the laying down of food standards as well as their harmonization with inter- national standards. Th is ministry is also the licensing authority for processed fruits and vegetable industries.

LABELLING ISSUES

India, along with a number of other countries, has supported the mandatory labeling of GM food by Codex.

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Limitations of the Regulatory System in India

I.Lack of adequate standards for risk assessment

II.The procedure and information required for a full environment

risk assessment and appropriate safety and emergency responses not

laid down.

III.Instructions and conditions for use on labeling and packaging of

products containing GMOs yet to be specified.

IV.Detailed safeguards as embodied in the Cartagena Protocol yet to

be incorporated.

V.Lack of infrastructure for risk assessment.

VI.Shortage of skilled personnel from laboratory researchers to

extension service officers. 34

(vii) Although food safety studies are fairly rigorous as prescribed by the RCGM, there are always uncertainties in the risk assessment process because this is a novel technology. Also there needs to be special considerations when the food forms a major portion of the diet (for e.g. cereals) and the special sociocultural habits of the population need the consumption of parts of the plant which are unconventional for other civilisations (e.g mustard leaves). In such a scenario special studies may have to be undertaken even on human volunteers.

Limitations contd…

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Presentation by: Guru P N 36