Quality Assessment and Maintenance of Genetically Altered and Specialised Seeds.

Why Specialised Seeds? Food provides lifeline of the people which derived from a variety of sources. The main story is agriculture. Successful agriculture largely depends upon a gamut of impulse in which seed plays key role. Emerging challenges are looming large for our survival and perpetuation at the interface of dwindling recourses. Population influx and steep competition for food, space and other basic amenities will be intensified considerably. By 2030, about 8 billion people will populate the world and by 2050, it is estimated to be 11 billion. Production of global staple food crops requires substantial increases in productivity without further increase in area covered with already intensively cultivated land. Expansion of area is only possible in regions having as marginal soils which is meager. The enormity of the challenge is significantly

increasing by decelerated availability of water and requirement of additional food resulting in steep rise in hunger, malnutrition and finally poverty across the globe. This is especially discernible in Afro-Asian countries evident in the recent past. There are numerous ways to increase agricultural productivity at sustainable scale. Dissemination of new knowledge and new technology along with the capacity to use it world over is the need of the hour. Modern agriculture has introduced large scale use of fertilizers, herbicides, pesticides and insecticides. Further, use of biological fertilizers, improved pest control, better soil and water management

conservation, and the use of improved seeds, produced by either traditional or biotechnological options are the other means for estimated yield enhancement. Seed comprising all forms of propagules, is considered to be the most vital input for enhanced productivity. A sea change has been witnessed towards better rural economy due to use of quality seeds of improved varieties in the recent past. In India despite of operation of several national projects like the National Seed Project since 1979-80, Annual

Oilseeds Scheme since 2004, Poverty alleviation in tribal areas through quality seeds (2004-2006), and latest Seed production in agricultural crops and fisheries since 20052006, there big gap still lies between the demand and production of quality seeds. Quality seed can enhance productivity upto 15-20% alone in diverse crops at marginal cost and when it is from a transgenic variety, it may constitute even more under optimal management. Genetically altered seeds, developed by recombinant DNA (r-DNA) technology is a scale neutral technology and could easily be adapted by the farmers[1]. GM technologies are relevant in the current agricultural scenario in India because not only it can take care of further environmental damage but also augment agricultural production and productivity, when properly integrated into traditional systems. Small and marginal farmers can also be benefited from the r-DNA technology when such genetically modified seeds become available to them at reasonable cost. It also has the potential to generate income at the local level among land less labour and to the people living below poverty line by extending the food security to grass root level.

Biotechnology is the applied part of to medicine and agricultural technology used in plant genetic improvement precisely for novel attributes. It is often called as genetic modification technology (GM technology) and was first developed during 1970s. Development of transgenic crops or seeds, like conventional plant breeding and farmers selection in the field, aimed at incorporating, removal or alternation of character(s) of choice in plants. It offers immense possibility of combining genes from different organisms or bringing desirable characteristics from related or unrelated species subsequently the product becomes a partner for use in conventional breeding. Modification in food crops by GM technology help improve human health by redressal of malnutrition, under-nutrition and is a route to reduce the national expenditure and resources required to implement the food supplementation programme. Other benefits from transgenic plants under study include increased flexibility in crop management, decreased dependency on chemical insecticides and soil disturbance, enhanced yield,

easier harvesting and higher proportions of the propagules available for trading. For the consumer this should be made available at low cost of food and higher nutritive value. Apomictic seeds, another form of specialised seeds, can arise from a plant's sexual cells, which do not enter meiosis required for sexual reproduction. Alternatively, seeds can be generated from non-sexual (somatic) cells. Sometimes, both sexual and asexual seeds develop from the same flower. Apomictic plants produce cloned seed, enabling them to reproduce asexually. But pollen being viable many a time, apomixis can also is transmitted through the more common mechanism of sexual reproduction. Apomixis was defined by Winkler as replacement of the normal sexual reproduction by asexual reproduction, without fertilization[2]. In nature, apomixis is widespread but infrequent occurs 10% of the 400 families of flowering plants. The most obvious benefit of introducing apomixis into crops would allow selecting an individual plant and propagate it as clones through its seeds. A second benefit would be to expand the range of wild relatives that could be integrated into breeding programmes. This is because asexual seeds can contain two sets of chromosomes of different sizes and still be viable, while equivalent sexual seeds would probably not develop. Apomictic seeds would demand dependency on fertilizers and pesticides. Like other hybrids, apomictic hybrids would still be designed to perform their best under certain environmental conditions, which small farmers are unlikely to achieve and maintain. Apomixis might increase farmers' access to hybrids, but not their control of them. Moreover apomictic seeds would allow farmers to fix the genetic characteristics of any of their individual crop plants, by crossing them with an apomictic line Apomixis would allow farmers to become faster breeders, just as it would for formal breeders. It would give farmers more control of their local agro-environment. It would theoretically guarantee yield and uniformity (and therefore, marketability) of their own selected varieties. Seeing its vast potentiality, production of artificial seeds has unraveled new vistas in plant biotechnology. The artificial seed technology is an exciting and rapidly growing area of research in plant cell and tissue culture. The idea of artificial

seeds was first conceived by Murashige[3] which was subsequently developed by several investigators. Initially, the development of artificial seeds had been restricted to encapsulation of somatic embryos in a protective jelly like sodium alginate etc. It had been considered that the induction of somatic embryogenesis (SE) and/or pollen embryogenesis which genetically differs from zygotic embryogenesis, is the prerequisite for the preparation of artificial seeds. Their induction has been reported in a number of cereals, millets, tuberous plants, vegetables, and other commercially important plants like soybean, mustard, coffee, tobacco, and cotton. However, because of certain inherent problems, the rate of production of uniform and high quality embryos is much lower as a result of which the preparation of efficient and quality seeds has been found to be successful in only a few crop plants like carrot[4] and alfalfa[5]. But artificial seeds are considered to be a very effective method for short and long term conservation of genetic resources through encapsulation and cryopreservation methodologies.

Quality control of specialised seeds

Genetic purity is the most important and essential criterion of any crop variety or its hybrids for its use in full potential. Grow-out test (GOT) and detection of off types is a popular and very often used methodology practiced by the researchers in detecting genetic purity. However, it is land and extensive labour intensive, time consuming tedious procedure involving high cost. Therefore, use of other methodologies came in-practice. With spectacular advancement in biotechnological

research and development, improved diagnostic testing equipment suitable for determining grain composition and purity are now available and in vouge. In all countries seed and grain purity is the main focal point in producing a specific crop with good quality and it is important that testing determines the level of purity. An Enzyme Linked Immunosorbant Assay (ELISA)[6] strip test is used to identify genetic traits in seed, leaf or grain. Strip tests

provide a simple Yes or No result within 5 minutes and if the result shows a negative a second test is conducted. The strip test works by identifying herbicide or pesticide traits within a sample and can deliver a pass or fail answer within minutes. The ELISA strip test is used as a guide to validate materials for audit or contract process with a Polymerase Chain Reaction (PCR) tests being the more acceptable standard technique. Companies exporting to countries requiring specific non-GM seeds opted for the more rigorous PCR test. The ELISA strip test is considered to be an acceptable field test and works well to maintain grain system integrity from the beginning. The strip tests are used to ensure methods of segregation are correct and lots are not mixed or adulterated. Countries like England and Ireland receive imported materials from countries that are known for growing GM crops. PCR tests are the accepted standards for contracts with represented samples from each shipment submitted for testing. The cost of PCR testing and the delays in receiving results are areas affecting importers substantially today. PCR testing does not pick up conventional soybean types. A concern is that there are presently no uniform international standards for PCR testing. However, in recent times PCR based technology is well accepted and emerged as very important to determine GM seeds through probe/primer based technology using Northern/Southern/Microarray hybridization technique. Moreover, purity of specialised seeds may also be tasted by using standard molecular markers such as: SSR, ISSR RAPD, AFLP, SSR etc. If properly used, these markers may assess genetic purity of any seed stock accurately precisely and with confidence. SSR markers can distinguish the high yielding variety from parental lines very perfectly. Through multiplex PCR, SSR marker combinations that were unique to a particular parental line or hybrid were also identified. Also introduction of parental and hybrid specific SSR markers helps to utilize the informative SSR markers for detection of contaminants in a cost effective manner. ISSR primers are dominant but are highly repeatable and deemed to be detecting number of alleles and appeared as a good choice as genetic marker. RAPD primers are dominant, technically simple to perform and less costly. Conversion of RAPD bands into SCAR

could lead to repeatability via generation of PCR specific primers and thus would be a good option for use in genetic purity testing. Therefore, the above mentioned molecular markers especially microsatellite (STMS/SSRs) and ISSRs are being used in the present study for genetic purity analysis of germplasm lines from specialised/genetically altered seeds obtained from commercially important crops including their hybrids. The Near Infrared Reflectance (NIR) machine is now being developed to measure herbicide and pesticide traits along with moisture, protein, oils, starch and fiber content. Grain harvesting machinery has been modified with an NIR machine to monitor crops coming off the field. Prototypes are still very much in their infancy development stage. These precise analytical instruments will not only be developed for laboratories but in future will be included as part of harvesting equipment. A continuous sampling process could be used to analyze and segregate crops directly off the field before delivery to the elevators.

Preparedness for Biosafety concerns

Biosafety is an essentially important aspect for genetically modified seeds. Biosafety by definition protects human and animal health and environment from the possible adverse effects of the product of modern biotechnology[7]. GMOs and r-DNA products in India are governed by the Environment Protection Act (EPA, 1986) which encompasses a frame work for the protection and improvement of environment provides a frame work for the protection and improvement of environment. Other rules related to transgenic seeds and plants include Industries (Development and Regulation) Act, (1951); Seed Act (1966); Seed Rules (1968); Seeds (Control) Order (1983); Seeds Policy (1988, 2002); rules on GMOs (effective from 1993) and Protection of Plant varieties and Farmers rights Act (2001). DBT along with other government organizations (viz, ICAR) formed collaborative frame work for monitoring evaluation and clearance of GMOs for commercialization. An interrelated mechanism

between different committees and developments of Government of India is instrumental in governing and regulating the works related to GMOs. (Table 1.)

Table 1: Various monitoring committees related to management of GMOs in India. COMMITTEE FUNCTIONS Recombinant DNA Advisory committee i) Review developments in Biotechnology (RDAC) at national and international levels. ii) Recommend suitable and appropriate safety regulations for India in rDNA research, use and applications. Review Committee on Genetic manipulation i) To note, approve, recommend (RCGM) generation of appropriate biosafety and agronomic data. Institution Biosafety Committee (IBSC) i) To note, approve, recommend and seek approval of RCGM Genetic Engineering Approval i) To approve large scale use Committee(GEAC) i) To approve open release to environment, inform decision to administrative ministry and applications /investigators to follow PVP/Seeds Act. State Biotechnology Co-Ordination i) Empowered to inspect, investigate and Committee (SBCC) to take punitive action in case of violations of statutory provisions through the State pollution control board or the directorate of health etc. ii) To review periodically the safety and control measures in various institutions handling GMOs iii) To act as nodal agency as DLC. District Level Committee (DLC) i) To monitor the safety regulations in installations inspect, investigate and report to SBCC/GEAC about compliance/ noncompliance of r-DNA guidelines or violation under EPA. ii) To act as a nodal agency at district level to assess the damage, if any due to release of GMOs and to take on site control measures

IPR issues The proprietary rights over knowledge and products in agriculture are getting increasing importance. These inputs are required to be protected through some form of Intellectual Property Rights (IPR) or else severe controversies for its proprieties might arise. US patents are even granted to plant and bacterial gene sequences, animal inventions and human genome sequences. There are cases of granting broad species patents in cotton and soybean. The proprietary rights granted to public or private firms are often justified showing the high costs of research, development with and biotech

commercialization

associated

inventions and to safeguard the inventions from piracy. According to section 3(j) of the Patents Act, plants and animals in whole or any part thereof other than microorganisms but including seeds varieties and essentially biological processes for production or propagation of plants and animals for example, clones and plant varieties are not patentable. However, processes leading to the development of Genetically Modified Organisms (GMO) can constitute patentable subject matter. The IP Protection of plant varieties including the extant varieties will be secured under the PPVandFR Act. (2001) As per the PPVandFR Act, an extant variety is the variety available in India which is either (i) notified under section 5 of the Seeds Act, 1966(54 of 1996), or (ii) a farmers variety, (iii) a variety about which there is common knowledge, or (iv) any other variety which is in public domain. For registration and protection of new varieties/hybrids, the principal investigator (PI)/plant breeder will inform the respective Institute Technology Management Committee/Zonal Institute Technology Management Committee at the institution about the availability of any prospective material developed by him/her, which can quality for a new, distinctive, uniform and stable crop variety as per the requirements of the PPV and FR Act. The IP rights in ICAR (Indian Council of Agricultural Research) are being followed by using various Indian Acts. These are as follows: The copyright Act, 1957 (last amended in 2000); The Patents Act, 1957 (last amended 2005); The Trade Marks

Act, 1999; The Designs Act, 2000 (last amended in 2001); The Geographical Indications of Goods (Registration and Protection) Act, 1999 (last amended in 2002); The Semiconductor Integrated Circuits Layout Design Act, 2000 (last amended in 2001) and the Protection of Plant Varieties and Farmers Right Act, 2001 (last amended in 2003). ICAR patent guidelines web sites are available at www.icar.org.in and

www.patentoffice.nic.in. Central Technology Management Committee has been constituted at the ICAR Headquarters under the chairmanship of the Director General, ICAR, which is the highest decision making body in the ICAR for IP management and technology transfer/commercialization. Agro-technology Management centers have also been established at ICAR Headquarters to manage its IPR portfolio and commercialize its IPR enabled technologies. Zonal Agro-Technology Management Center is the center established at the identified zone/ institute to manage IP protection/maintenance and technology transfer/commercialization of the institute. Zonal Institute Technology Management Committee involves the committee constituted at the identified zonal institute to facilitate/ manage IP protection/maintenance and technology

transfer/commercialization for ICAR institutions in the zone. Institute Technology Management Committee has constituted at the level of an ICAR institution chaired by the Institute Director, for addressing IP related matters of the institution. Exclusive license of an IPR enable technology in ICAR means a license, which will entitle the licensee to exclude all other persons (including the patent holder) in the commercial use of the technology covered in the licensee. In licensing means acquiring research-tools that are already protected by patents/IPR for research and technology generation under specific terms and conditions, e.g. research/commercial use. License-out means the document embodying legal permission from ICAR to the other party to use its technologies/IP/knowledge for commercial or other purposes under the terms and conditions and limitation, including a license fee and/or royalty, as negotiated and specified in the license. Non-exclusive license of an IPR enabled technology donates a license, which would confer on the license the right to commercially use that technology whereas, at the same time the same right could be also be made available to other licensee(s) on same, similar or with different terms and conditions.

Controversies and ethical issues

Most of the controversies in transgenic seed production arise either for reduced production costs in agricultural areas that already have high productivity levels for value addition to the final product. There has been much debate about the potential benefits and risks that may result from the use of transgenic crops. Creation of genetically modified organisms (GMOs) is often presented as a magic solution or universal panacea for the problems of poverty, inadequate food access, nutrition and environment degradation across the world. Conversely, there are people who display GM crops as the Frankensteinian monsters and appeared human health hazards, which interfere rapidly and too intensively with natural processes of transgenic evolution[8]. A basic question, of course, is whether the new GM technology is safe, and this is absolutely crucial since the effects may only be known much later. The critics of this technology argue that in any new technology, it is always possible that harmful side effects may occur, and therefore there need to be long term tests on health and environment before its commercial cultivation. Similarly, unlike traditional plant breeding methods, the new technology uses artificial laboratory techniques to combine genes that would never occur in nature, which really means altering genetic patterns that have developed over millions of years. There are possibilities, especially for the third world that they might face the menace of pollen transfer, appearance of insecticide resistance super-weeds, virus resistance and other genetically determined traits, with possible food chain and weed menace consequences. The process of globalization of the food system and marketing and the extension of industrial patenting and other intellectual property systems to living organisms further, led to the widespread cultivation and rearing of fewer varieties and breeds for a uniform but competitive global market. In nutshell, GM seeds with wonderful properties will get overall acceptance if managed with more consciousness and handled with proper care following the guidelines stringently as outlined by various committees and research for safe handling of GM crops.

Reference: 1. Datta, S. K. and Potrykus, I. 1989, Theor. Appl. Genet., 77: 820824. 2. Frontline 2001 vol 18-Issue 12 Jun. 09-22, 3. Fujii, J, A. Slade Redenbough, K 1989, In vitro Cell Dev. Biol., 25: 1179. 4. Luytz Grohmann, 2009 Biotechnology in Agriculture and Forestry: Genetic Modification Of Plants: 64 (7) Springer 15-12,. 5. Murashige, T 1978, In Frontiers of Plant Tissue Culture: Thorpe, T. A., 15. 6. Ramanaiah T.V. 2006 Short-term orientation course on biosafety and biotech regulation 6 -11 Feb, at TERI, New Delhi. 7. Tuli Rakesh, Bhatia C. R., Singh P. K. and Chaturvedi Ratnesh 2000, Curr. Sci 79, NO. 2, 25 8. Winkler, H. 1908. ber Parthenogenesis und Apogamie im Pflanzenreich. 2 (3) Progressus Rei Botanicae 293454.