Rajasekharan P.E.

Research highlights In the last 25 years technology has been developed for long-term cryopreservation of Nuclear Genetic diversity. Long term cryopreserved pollen in the form of nuclear genetic diversity (NGD) of citrus, papaya, grape, mango, tomato, eggplant, onion, capsicum, rose, gladiolus, gerbera, carnation and RET species of medicinal plants are maintained in liquid nitrogen. The pollen cryobank is managed by periodic replenishment of the cryogen, for maintaining a constant cryogenic temperature throughout the storage duration. This technology developed is globally acceptable and will be useful for production of hybrid seeds in the above-mentioned crops, besides its use in gene banks. At the pollen cryobank of IIHR, information pertaining to 650 pollen samples of more than 45 species belonging to 15 families is stored using this database. The software support provided for this database was FoxPro version –2. For easy management of data one main database and three sub-databases were created. Genus, species and cultivars information is available in a separate database. Information pertaining to longevity, viability and fertility of pollen after cryopreservation, media used for pollen germination, year wise listing of collected samples, year-wise list of samples cryopreserved in different years are available within the database, which is menu driven and user friendly. Although genetic conservation through pollen storage does not accomplish whole genome conservation, a breeder involved in genetic enhancement of a given crop can have access to a Pollen Cryobank facility, for nuclear genetic diversity (NGD) inputs in his amelioration program. Besides the already existing role of pollen cryobanks in breeding, there are many promising applications, which has come to focus with the recent advances in allied bio-scientific areas. THE POLLEN CRYOBANK AT IIHR, BANGLORE FINDS MENTION IN LIMCA BOOK OF RECORDS-2001. The pollen samples belonging to the following species were stored in liquid nitrogen. A total of 660 samples are there at present Effect of electromagnetism on pollen germination and viability: The effect of electromagnetism on pollen germination and vigour was studied using balsam pollen. Positive results were obtained for increase in germination profiles and vigour. The pulsed medium was also shown increase in germination and vigour in balsam pollen From 2004 onwards 1.1.2 Sustainable Management of Vegetable and Ornamental Plant Genetic Resources (Project leader) 1.1.1 Sustainable Management of fruit and medicinal Plant Genetic Resources (Project Associate) Research highlights A total of 14-exploration and collection missions in the 3 year period were carried out under this project in hotspot areas of south India. The germplasm were collected along with the passport data Externally aided projects 1)FAO sponsored Hortivar Project team member 2)Co-ordinated IIHR botanical garden activities for Botanical Garden project of Botanic Garden Conservation International (Coordinated by NBRI, Lucknow) 3)In vitro conservation of grape germplasm In the ICAR ad-hoc research project “In vitro conservation of Grape germplasm and establishment of in vitro Active Gene Bank at IIHR” Bangalore, acted as the Co-Principal Investigator 1997-1999. With the support of an ICAR Ad-hoc Scheme, protocols for conserving grape genetic resources was developed at IIHR for accessions acclimatized to Indian ecosystems. It is now possible to maintain grape vitroplants for periods ranging from 6 months to 1.5 years at low temperature without an intervening sub-culture. In vitro conservation repositories involved in conservation of grape PGRs can straight away use this technology developed. 4)National Agricultural Technology Project of NBPGR on Plant Biodiversity NATP project of NBPGR on Plant Biodiversity as a Co-CCPI for carrying out exploration, collection, domestication evaluation and conservation of endangered medicinal plant species of south India. Carried out more than 18 plant exploration missions in various parts of south India and collected many species of economic importance (1999-2004) Gist of work done Successfully completed research in the area of Plant Biodiversity carried out exploration missions in the forest ecosystems under the National Agricultural Technology Project, dealing with exploration, collection, evaluation, characterization and conservation of plant species native to Indian sub-continent. IIHR as a co-operating center of NBPGR has carried out 18 exploration missions in the Karnataka/ Kerala region So far thousand accessions of medicinal plants have been collected, comprising of live plants, seeds, cuttings, etc., a field gene bank has been established maintaining over 100 species, the collected accessions maintained as live plants. 37 accessions have been characterized from among the collections In vitro conservation protocols have been optimized for 25 species. 85 accessions in the form of seed, live plants, cuttings & in vitro plant material have been deposited with NBPGR. The program supported inventorization of indigenous plant diversity distributed in the sub-continent, besides attempting to identify potentially new species, which could be brought under cultivation mode. My self was actively involved in o Planning & execution of exploration and identification of plant species missions for collection of plant species from forest ecosystems o Domestication, Evaluation and Characterization, conservation and documentation of PGR o 32 of the 40 species listed as per priority have been collected from the exploration missions conducted. Strategies for exploration and collection, evaluation and conservation of vegetable and ornamental species devised and implemented in the ongoing project on Management of Horticultural Genetic Resources Established a field gene bank for medicinal plants The co-operating center has carried out 17 exploration missions in the Karnataka/Kerala region assigned under zone-II sub-zone-8. A total number of 970 accessions have been collected, comprising of live plants, seeds, cuttings, etc., A field gene bank has been established maintaining 100 species collected accessions as live plants. 20 unique accessions were collected from the 17 exploration missions carried out by our center. 32 species of the 40 species listed as per priority have been collected from the exploration missions conducted during 1999- 2003. 37 accessions have been characterized from among the collections In vitro conservation protocols have been optimized for 24 species 111 accessions in the form of seed, live plants, cuttings & in vitro plant material have been deposited with NBPGR. 5) Collection, chemical and molecular profiling, domestication and conservation of Mappia foetida an endangered medicinal plant GO/KR-12/2007 from National Medicinal Plants Board, New Delhi Research highlights: 1. Exploration and collection missions Eight explorations were carried out for collecting Mappia foetida in Shimoga and Yercaud in Salem. Near Shimoga a four-year plantation established by Sri N.G Pai in Sagar, under open field conditions and the growth of the plants was not satisfactory. All the plants were slow growing populations in the forest near Kodluthatte. Unlike the captive populations these natural population was healthy. Leaves were big in comparison with the captive population; all plants were growing under shade. An exploration and collection mission to Yercaud in Salem district, Tamil nadu was carried out during January 2008 to collect Mappia foetida. Sri. V. R. Puasalkar, of Aruna Planta Medica has established Mappia foetida in 100 acres Coffee plantations in Yercaud, a hill station 222 km from Bangalore. In both explorations root, shoot and leaf samples were collected for chemical and molecular analysis in additions to the seedlings. Exploration and collection missions to Agumbe, Joida, Amboli ghat, Khanapur and Jambotti, Sirsi, Wayanad, Dhakshina Kannada, Idukki District of Kerala, Coorg were carried out to collect seeds, materials for chemical and molecular profiling( leaves, root bark, stem bark etc). Seedlings were also collected. In situ evaluation of populations were also carried out to see how each population deferred from each other in terms of diversity in morphological trait Tissue culture systems: Efficient in vitro systems were established using embryos as explants. In vitro conservation using low temperature and low light was successful in Mappia and up to one year the cultures could be maintained at 10°C without subculture. Genetic fidelity of tissue cultured Mappia foetida seedlings were tested using ISSR markers. Mapping of Nothapodytes nimmoniana in Western Ghats Using the passport data the distribution of Nothapodytes nimmoniana species were mapped using DIVA GIS software. The results are the form of maps. Domain model DIVA GIS (Hijmans et al, 2003) was used to predict the potential distribution of Nothapodytes nimmoniana in Western Ghats. BIOCLIM climate parameters were used for this study. The distribution of the N. nimmoniana species was mapped using the primary data which obtained from the various exploration and collection missions. The latitude, longitude and altitude of the locations were obtained using a Global Positioning System. The geographical coordinates from the primary data set (n=75) was digitalized. Prediction of species distribution patterns With the help of Bioclim /Domain tool extracted the data from the climate database for point locations and predicted the possible occurrence of N. nimmoniana FGB: A field gene bank was also established for Mappia foetida with the accessions collected during the exploration and collection missions from Western Ghats 6) Acted as Principal Investigator of 2.37 Crore Network project on RET medicinal plants ex situ conservation and utilization in South and North East India Gist of work done IIHR as a lead center in the network project on RET medicinal plants , sponsored by NMPB, exploration and collection missions were undertaken in Kerala, Karnataka and Tamil Nadu. Out of 31 collection missions-374 accessions collected The collected accessions were brought to Bangalore and established ex situ in Field gene bank for further studies. Mapped areas of distribution and predicted the possible areas of distribution- for 32 targeted species using DIVA GIS software A seed bank was also established for Oroxylum indicum(140), Decalepis hamiltonii(6), Holostemma adakodien(3) Embelia tsjeriamcottam(3), Eemeblia ribes (1). maintaining Accessions from diverse locations. Voucher samples were deposited in national gene bank of NBPGR and IC Nos obtained – for 32 seed propagated species. Accessions of Kaempferia galanga(11) Kaempferia galanga( 8), Holostemma adakodien (4), Oroxylum indicum (9), Embelia ribes (2), Celastrus paniculatus (23), Decalepis hamiltonii (6), Aristolochia tagala (3), Saraca asoca (6), Alpinia galanga19,Kaempferia galanga 26,Rauvolfia serpentina10 could be established in the FGB of IIHR. In vitro conservation In vitro multiplication and conservation protocols were optimised for Alpinia calcarata, Kaempferia galanga and Acorus calamus.. Seed: Seed cryopreservation were initiated with the following RET medicinal plant species. 1. Oroxylum indicum 2. Operculina turpethum 3. Holostemma adakodien 4. Celastrus paniculatus 5. Decalepis hamiltonii Documentation A database for these 32 selected RET Medicinal Plants. Visual basic 6.0 and MS access 2003 was used for developing this database which is named as “RET Medicinal plant Database”( RETMED base). MS Access is a powerful RDBMS. The passport data, evaluation data (morphological chemical and molecular) were entered in MS. Access. Visual Basic 6.0 is simple as well as user friendly and was used as the front end ie. as a user interface and MS Access was used as the back end, ie: as a data storage part. Database connection is established by using ADO data control, one of the ActiveX control used for data access from databases to VB 6.0. Coding was done in such a way that, the user can access all the data, generated in RET Medicinal plant species from a single window. In the database home window consist of menu buttons for accessing passport data, evaluation data and abstracts of published research papers different RET medicinal plant species. The distribution and prediction maps generated through DIVA GIS software for each species were also incorporated in the database. Other than this the database includes the photos of each RET Medicinal plant(habit and habitat, flower, fruit seed and economical parts) data from exploration, field gene bank, in vitro studies , anatomical studies. Pollen studies were also incorporated . Molecular characterization : Molecular characterisation of ,Acorus calamus, Alpinia galang, Alpinia calcarata,Kaempferia galanga, Decalepis hamiltonii, Orxoxylum indicuam and Mappia foetida from different locations(western ghats and North east region) were subjected to ISSR analysis. Chemical characterization Decalepis hamiltonii , Oroxylum indicum , Celastrus paniculatus , Alpinia galanga and Kaempferia galanga chemically profiled Research at Biotechnology laboratory of Central Plantation Crops Research Institute Kasaragod ,Kerala Clonal propagation of Oil Palm (Elaeis guineensis Jacq),coconut ,cardamom and cashew carried out for about four years as Senior research Associate(from 28-05-1985 to 03-09-1986) and Technical Assistant T-II-3(03-09-1989).During that period I was involved in the in vitro conservation of coconut embryos by using minimal medium and low temperature. After joining as scientist (06-06-1989) I underwent a core training programme at National Bureau of Plant Genetic Resources (NBPGR), New Delhi for six months from 7-11-1989 to 27-4-1990.In that training programme, various aspects of Plant genetic resources activities like collection, evaluation and conservation of plant germplasm components such as seed, pollen, in vitro material under low temperature and cryogenic condition iii. Technologies developed / Technological contributions • Technology has been developed for long-term cryopreservation of Nuclear Genetic diversity. Long term cryopreserved pollen in the form of nuclear genetic diversity (NGD) of citrus, papaya, grape, mango, tomato, eggplant, onion, capsicum, rose, gladiolus, gerbera, carnation and ret species of medicinal plants are maintained in liquid nitrogen. The pollen cryobank is managed by periodic replenishment of the cryogen, for maintaining a constant cryogenic temperature throughout the storage duration. This technology developed is globally acceptable and will be useful for production of hybrid seeds in the above-mentioned crops, besides its use in gene banks. IT FINDS MENTION IN LIMCA BOOK OF RECORDS-2001 • Effect of electromagnetism on pollen germination and tube growth studied in detail • Effective pollen collection, germination protocols developed for a wide array of species of horticultural importance (vegetable, ornamental, fruits and medicinal species • First time in the world mango pollen was cryopreserved and pollination with the same resulted in fruit and seed set • Character transfer using cryopreserved pollen was established for the first time using cryopreserved trifoliate orange pollen • Pollen of wild relatives of capsicum, tomato and solanum were cryopreserved for the first time and used in the pollination with cultivated species resulting in fruit and seed set • Exhaustive studies on gametophytic selection using pollen for heat tolerance in vegetable crops viz., tomato, brinjal and caspsicum and papaya • Protocols for in vitro culture optimization, multiplication and in vitro conservation of Carnation • Protocols for in vitro culture optimization, multiplication and in vitro conservation of Chrysanthemum • Protocols for in vitro multiplication and conservation for jack fruit • Protocols In vitro system for jack using embryonic axes • Design establishment of Field Gene Bank for RET medicinal plants • Protocols for Nothapodytes nimmoniana Tissue culture systems and in vitro conservation • Distribution mapping of RET medicinal plants POLBASE- DIGITIZATION OF INFORMATION MANAGEMENT SYSTEM FOR POLLEN CRYOBANKS was developed. At the pollen cryobank of IIHR, information pertaining to 600 pollen samples of more than 45 species belonging to 15 families is stored using this database. The software support provided for this database was FoxPro version–2. For easy management of data one main database and three sub-databases were created. Genus, species and cultivar information is available in a separate database. Information pertaining to longevity, viability and fertility of pollen after cryopreservation, media used for pollen germination, year wise listing of collected samples, year-wise list of samples cryopreserved in different years are available within the database, which is menu driven and user friendly