RECENT AGRICULTURAL DEVELOPMENTS IN JUTE, KENAF AND MESTA THROUGH TRADITIONAL AND BIOTECHNOLOGICAL APPROACHES

Dr. Asma Khatun Chief Scientific Officer Genetic Resources and Seed Division Bangladesh Jute Research Institute Manik Mia Avenue, Dhaka 1207, Bangladesh Abstract In Bangladesh, the major part of agricultural researches on jute, kenaf and mesta are being

conducted in Bangladesh Jute Research Institute (BJRI). BJRI has been conducting researches in agricultural aspects with the objectives to develop new varieties of jute, kenaf and mesta with higher yield, better fibre quality, photo neutrality, short field duration, water logging tolerance, pest and disease tolerance. Other practices are crop rotation, ribbon retting, seed purification through botanical, late jute seed production and jute, kenaf & mesta germplasm collection and their characterization. Since inception BJRI has developed 36 varieties of jute, kenaf and mesta of which 12 varieties have already been released for commercial cultivation. Strains have been developed with super white fibre having least cuttings. Leaf mosaic resistant jute, nematode resistant kenaf and less photosensitive and early maturing mesta strains have been developed. Ribbon retting technique in water deficit areas is a remarkable achievement. The Central Research Institute for Jute & Allied Fibres (CRIJAF) in India deals with white and tossa jute, mesta and kenaf CRIJAF, have yielded a number of varieties suitable for various situations and purposes. Early tossa and white jute varieties were developed. Varieties were also developed with higher yield and responsive to higher level of nitrogenous fertilizer. A lignified phloem fibre deficient (dlpf) mutant of jute has been identified. Researches are going on with jute based cropping sequence. CRIJAF has identified a variety of mesta which yields around 50-60 tons of bio-mass per hectare. A high yielding variety was developed with resistant to drought and lodging. Researches are going on in the USA on different aspects of kenaf production e.g. variety selection, the yield, resistance to diseases, nematodes, pests and weed control. Growers and researchers of the U.S. have found kenaf to be resistant to most plant diseases. The variety development work done by the ARS in Florida was very effective in breeding and selecting for anthracnose resistance. There has been little economic damage to kenaf by insects in experimental production fields in U.S.A. In the field of biotechnology, some developments have been made on jute, kenaf and mesta. Isolation of protoplasts, somatic hybridisation and genetic transformation have been attempted in Bangladesh and India. Protocols for plant regeneration from jute (C. capsularis and C. olitorius), kenaf and mesta have been developed as it is a prerequisite of transformation. DNA fingerprinting of 18 jute genotypes composed of popular varieties and some promising lines from both C. olitorius and C. capsularis have been done using RAPD and SSR techniques. A molecular marker for low temperature tolerance was developed. RT-PCR was conducted using the total mRNA isolated from three jute strains. 1

Introduction Jute, kenaf, and mesta are the second most important natural fibers next to cotton. Jute grows under wide variation of climatic conditions and stress of tropic and subtropics. It is grown in Bangladesh, India, Myanmar, Nepal, China, Taiwan, Thailand, Vietnam, Cambodia, Brazil and some other countries. Jute and kenaf are cultivated almost exclusively in developing countries of East Asia and in some parts of Latin America. Bangladesh, India and Thailand account for over 90 percent of world production. Jute production was 2.09 million tons, among which, production from Bangladesh accounted for 68%, India for 30% and Myanmar and Nepal for 1% each. Total kenaf production in 1999/2000 was 0.51 million tons, among which production from China accounted for 44%, India for 39%, and Thailand for 12%. Of the total world jute production, five producing countries, namely Bangladesh, China, India, Nepal and Thailand account for about 95%. These countries also account for 90% of export of jute products. In Bangladesh, the major part of agricultural researches on jute, kenaf and mesta are being conducted in Bangladesh Jute Research Institute (BJRI). Bangladesh Jute Research Institute (BJRI) is the oldest research institute of the country and has been conducting researches in agricultural aspects on two bast fibre producing species of jute namely, Corchorus olitorius (tossa jute) and C. capsularis (deshi jute) and two species of Hibiscus namely. H. cannabinus (kenaf) and H. sabdariffa (mesta / roselle). The major agricultural research priorities of BJRI are:

Development of HYVs for the favorable ecosystem under changing socio-economic situation with short duration, early sowing, temperature tolerance with achievable yield of 3-4 tons/ha including biomass of 6-8 tons/ha. Development of varieties of jute, kenaf and mesta having high biomass and suitable for year round cultivation for producing pulp for paper. Development of new varieties of jute for uses as vegetables. Production and maintenance of genetic purity of breeder seeds of different released varieties of jute, kenaf and mesta. Establishment of methods for transferring useful genes. Ascertaining appropriate technologies to reduce cost of production including labor cost. Soil and fertilizer management on jute and allied crops for paper pulp production in mill areas. Participatory jute farming systems research. Design and development of low cost semi mechanized ribbon for water scarce jute growing areas. Development of dry jute ribbon retting technique.

Achievements of BJRI As the cultivated lands are reduced because of the growing demand of food crops, jute and allied fibres are pushed to the marginal lands. Still the average yield of the country becomes steady. Development of photo-insensitive tossa varieties has increased cultivation area of tossa varieties which is now 60 (tossa): 40 (deshi) instead of 40: 60. The following technologies have been developed by BJRI.

Development of high yielding jute, kenaf and mesta varieties Since inception BJRI has developed 36 varieties of jute, kenaf and mesta of which 12 varieties have already been released for commercial cultivation. Out of 12 varieties, 5 varieties for deshi jute, 4 varieties for tossa jute, 2 varieties for kenaf and one variety for mesta. These varieties are cultivated through out the country in large scale. The national production of jute is 1.93 ton/hectare. On the other hand, production of BJRI evolved varieties are 2.50 Ton /ha. If jute can be cultivated with proper manage and care average yield could be increased. A short description of 12 released varieties is given below. A. The following Deshi jute varieties are presently cultivated throughout Bangladesh

Name of the varieties Agronomic characteristics of the varieties and releasing time BJRI deshi -2 High yielding variety. Very popular. Sowing time : 30 March-15 April. CVL-1 Maximum production 5.16 ton/ha and production at farmers field is 1977 2.46 Ton/ha. BJRI deshi -3 Early maturing type. Quick growing. Suitable for 3 crop rotation. CVE-3 Sowing time : 30 March-15 April. Maximum yield 4.51 Ton/Ha and 1977 average yield at farmers field is 1.97 Ton/Ha. BJRI deshi -4 Suitable for early sowing. Quick growing. Suitable for 3 crop rotation. CC-45 Sowing time : 3 February-15 April. Maximum yield 5.16 Ton/Ha and 1979 average yield at farmers field is 2.49 Ton/Ha. BJRI deshi -5 Short day tolerant type. Suitable for early sowing. Quick growing. BJC-7370 Suitable for 3 crop rotation. Sowing time : 15 March-15 April. 1995 Maximum yield 5.46 Ton/Ha and average yield at farmers field is 2.45Ton/Ha. BJRI deshi -6 Quick growing and early maturing type. on. Sowing time : 30 March BJC-83 15 April. Maximum yield 4.72/Ha and average yield at farmers field is 1995 212 T/Ha. B. The following Tossa jute varieties are presently cultivated throughout Bangladesh BJRI Tossa-1 O-4 1967 BJRI Tossa-2 O-9897 1987 BJRI Tossa-3 OM-1 1995 BJRI Tossa-4 O-72 2002 High yielding. Late maturing type. Can be sown in high land. Proper sowing time: 15 April to 15 May. Maximum yield 4.51 Ton/Ha and average yield at farmers field 2.32 Ton/Ha. High yielding. Can be sown early. Proper sowing time: 30 March 30 April. Maximum yield 4.61 Ton/Ha and average yield at farmers field 2.73 Ton/Ha High yielding. Can be sown early. Les photo sensitivity. Proper sowing time: 20 March to 30 April. Maximum yield 4.62 Ton/Ha and average yield at farmers field 2.49 Ton/Ha. High yielding. Can be sown early. Proper sowing time: 15 March to 30 April. Maximum yield 4.96 Ton/Ha and average yield at farmers field 2.90 Ton/Ha.

C. The following kenaf varieties are presently cultivated throughout Bangladesh BJRI Kenaf-1 HC-2 1977 High biomass production. Suitable for paper pulp production. Can be sown both in low and high land. Resistant to nematode. Proper sowing time: 15 March to 15 April. Maximum yield 4.63 Ton/Ha and average

BJRI Kenaf-2 HC-95 1995

yield at farmers field 2.60 Ton/Ha. High biomass production. Suitable for paper pulp production. Early maturing type. Can be sown both in low and high land. More resistant than HC-2 to nematode. Proper sowing time: 15 March to 15 April. Maximum yield 5.45 Ton/Ha and average yield at farmers field 2.70 Ton/Ha.

D. The following kenaf varieties are presently cultivated throughout Bangladesh BJRI Mesta- 1 HS-24 1977 High yielding. Resistant to nematode. Can be sown in sandy marginal lands. Proper sowing time: 15 March to 15 May. Maximum yield 4.70 Ton/Ha and average yield at farmers field 2.70 Ton/Ha.

Cultivation ratio of deshi and tossa jute Although tossa jute produces superior quality of fibres in comparison to deshi jute, the former cannot withstand water logging or inundation. Bangladesh is mainly constituted with low lying areas and is inundated by flood water. The ratio between deshi and tossa jute during 20s was about 80:30. Through giving adaptive selection pressure, the ratio was shifted to 70: 30 in 60s and after development of less photosensitive variety O-9897 and OM-1, the present ration between deshi and tossa is about 60:40. Jute based profitable crop rotation BJRI has developed profitable technology for three crops-based crop rotation. For example: jutelentil+mustard, jute-transplanted Aman-wheat, jute- transplanted Aman-gram, jute-transplanted Aman-onion, jute-transplanted Aman-vegetable, jute-transplanted Aman-mung bean or gram, jutetransplanted Aman-maize, jute- transplanted Aman- data, jute- transplanted Aman-gram+lentil, jute-transplanted Aman-potato+tobacco, jute- transplanted Aman-tobacco, jute-radish-wheat+lentil, jute-radish-potato+red spinach+pumpkin. The farmers have increased their income by using crop rotaion methods. Improvement of retting techniques The quality and quantity of jute production is very much dependant on proper retting. Improvement of retting technique through addition of urea and by removing the shamla colour of jute by washing in tamarind solution is one of the important innovations. Ribbon retting technique in water deficit area is a remarkable achievement. This technology is recommended for the area with water deficiency of water during retting season. Insect and pest control Beside recommendation of several pesticides, physical and biological control measures have also been recommended for jute. Raw garlic paste 12.50% i.e. 125 gram garlic paste mixed with 1 Kg jute seed can be used for seed purification and to prevent seedling mortality. Neem leaf extracts and water (1:20) can be used to control white mite. Fertilizer application Instead of using blanket fertilizer doses for all the varieties and locations, variety and soil specific fertilizer doses have also been prescribed. In jute crop, fallen leaves and left over roots after harvest enrich soil organic matter through degradation.

Development of late jute seed production technology To solve the problem of jute seed scarcity, BJRI has evolved a technology for late jute seed production. In this technique, jute seeds are sown in August-September instead of March-April. Seeds can be harvested in December. Around 500-600 Kg seed /Ha can be collected by using this technology. Top cutting of the jute plants for seed production was found to be another technology for jute seed production. Collection of germplasm of jute, kenaf and mesta There is a germplasm depository (Gene Bank) at Bangladesh Jute Research Institute (BJRI). This Gene Bank has a capacity of preserving one Lac accessions of germplasm with 200gm volume. At present, in the gene bank of Bangladesh Jute research Institute (BJRI) there is a collection of total of 5935 accessions preserved in two cells, one set at 20oC and another set is under +4oC. A total of 53 species of fibre producing plants are available. Of which, 15 species for Corchorus, 22 for Hibiscus and 16 species for other allied fibres of other genera are available. For Corchorus, 3344 accessions are of indigenous collection and 737 accessions were collected by International Jute Organization (IJO). For Hibiscus species, 585 BJRI collection and 925 are from IJO. Among the allied genera, one accession is from BJRI and 343 accessions are from IJO. About 3000 germplasm have been characterized. Development in India The Central Research Institute for Jute & Allied Fibres (CRIJAF) in India deals with white jute, tossa jute, mesta and kenaf CRIJAF, have yielded a number of varieties suitable for various situations and purposes. Five early tossa jute varieties were developed for sowing during middle of March without risk of premature flowering. Two white jute varieties have been developed which can be sown during early March with residual soil moisture which are very much suitable for low lying flood prone areas of N. Bengal and Assam. Shyamali is a white jute variety suitable for mid and high land and responsive to higher level of nitrogenous fertilizer. Among all the white jute varieties JRO 7447 offers the highest fibre yield. Two varieties are adapted to mid and high land situation and can be grown across the white jute belt of the country with a promise of high yield. Under rainfed situation sowing of jute is often delayed. Some JRO 632, JRO 3690, and JRO 66 are very much suitable for sowing during middle of April and gives remunerative yield. A lignified phloem fibre deficient (dlpf) mutant of jute has been identified. Results of jute based cropping sequence revealed that the rotation of Jute-Paddy-Tomato is found most profitable. Technology developed for extraction of bast fibre (Jute, Mesta, Sunnhemp and Ramie) with the help of machine. JRC 4444 (Baldev) is a variety suitable for Orissa region. It is a selection from a multiple cross and offers around 30 q/ha of fibre under optimum growing condition. Research achievements of CRIJAF Sustained efforts at Division of Crop Improvement, CRIJAF, have yielded a number of varieties suitable for various situations and purposes. They are described below: Tossa jute varieties for early sowing and multiple cropping system Five tossa jute varieties are available for sowing during middle of March without risk of premature flowering and can be harvested by middle of July facilitating transplantation of 'Aman' paddy on the same land. The varieties are JRO 524 (Navin), JRO 7835 (Basudev), JRO 878 (Chaitali tossa), JRO 8432(Shakti) and JRO 128(Surya). These varieties are very much suitable for multiple cropping based intensive farming system. Fibre quality of JRO 128 is found better.

White jute varieties for early sowing and low lying areas Two white jute varieties namely JRC 321 (Sonali) and Hybrid 'C' (Padma) have been developed which can be sown during early March with residual soil moisture. They are very much suitable for low lying flood prone areas of N. Bengal and Assam. Early sowing enables the varieties to be harvested early before inundation takes place. White jute variety with finer fibre for diversified products Furnishings, dress materials, floor coverings, wall hangings and other uses of jute are proliferating these days. For such uses jute variety with finer fibre is a need of the day. Fibre fineness of jute varieties released so far, ranges between 16-18 denier. However, JRC 321 produces fibre having fineness around 10 denier. Such fibre is now in great demand in the diversified jute sector. Efforts are going on to develop jute varieties having fibre finer than JRC 321. White jute variety responsive to high fertility level JRC 7447 or Shyamali is a white jute variety suitable for mid and high land and responsive to higher level of nitrogenous fertilizer. Among all the white jute varieties JRO 7447 offers the highest fibre yield under 60 kg. N/ha or above. White jute varieties for mid and high land with wide adaptability JRC 212 (Sabuj Sona) is an old variety developed by direct selection from an indigenous germplasm. Another new variety, JRC 698 (Shrabanti), released in 1999 was developed by making multiple-crosses. These two varieties are adapted to mid and high land situation and can be grown across the white jute belt of the country with a promise of high yield. White jute varieties for specific locations JRC 4444 (Baldev) is a variety suitable for Orissa region. It is a selection from a multiple cross and offers around 30 q/ha of fibre under optimum growing condition. Mesta varieties for high fibre yield

Several mesta varieties have been developed which are popular among the farmers. H.S. 4288 is a variety with green stem but red pigment at the nodes, bristled, deeply lobed lamina. It yields 18-20 q/ha. H.S. 7910 (Ujjal) is a non bristle variety with green stem but irregular flush of red pigment extending with maturity of crop, lamina deeply lobed, yields 18-20 q/ha resistant to drought and lodging.
Mesta variety for jam, jelly and sauce preparation Long and fleshy calyx mutant of mesta was identified at CRIJAF and used for preparation of jam, jelly and sauce. Food testing laboratories of Calcutta and Lucknow certified the products to be absolutely harmless for human consumption. Initial study has suggested it to be economically viable special features of those fleshly calyxes are: (i) Calices are rich in pectin, hence no additional preservatives is required to be added. (ii) No artificial colour is needed for the products as the calyces have deep red natural pigment. (iii) Calices become available in the month of December, January when neither guava nor mangoes are available.

Kenaf variety for high fibre yield Widely adapted kenaf variety H.C. 583 is available which gives high fibre yield with yield potentiality of 25-28 q/ha. This is a selection from an exotic germplasm of Nigeria. Kenaf variety for pulp and paper Division of Crop Improvement, CRIJAF has identified a variety of kenaf namely MT 150 which yields around 50-60 tons of bio-mass per hectare. News print made out of the whole kenaf plant, shorn off leaves, is better than imported Canadian and Russian newsprint. High yielding jute varieties in the pipe line There are several elite lines of jute, kenaf and mesta in the pipeline which have shown promise of higher yield over the existing varieties. Promising lines S 19 of tossa jute, CC 4, CC 15, and C 80 of white jute, JBS 10,NBr 4 and AMT of H. S. mesta and JBM-89-1306 of H. C. mesta developed at Division of Crop Improvement, CRIJAF, have been recommended for adaptive trials at farmers' field by All India Coordinated Project. Research Achievements Jute has been proved to be a C3 plant yet its radiation use efficiency is above 3.0 g MJ-1. Most of the assimilated carbon and energy remain trapped in the wood (xylem) that are not utilized adequately for the purpose of commerce. A lignified phloem fibre deficient (dlpf) mutant of jute has been identified and characterized to show that genetic regulation of bast fibre development may be more complex than had been presumed. The biochemical consequences of this mutation, namely an apparent decrease in lignin content and its effect on other constituent of the cell wall will have important implications for crop biotechnology Achievements Maximum yield of fibre was obtained from the various JRO-524 (29.28 q/ha) followed by JRO8432 (28.58 q/ha), S-18 (28.31 q/ha), S-19 (27.38 q/ha), JRO-66 (26.70 q/ha), CO-28 (26.57 q/ha) and PPO-4 (18.84 q/ha) during the year 2000 2003. Results of jute based cropping sequence revealed that the rotation of Jute-Paddy-Tomato is found most profitable on the basis of C:B ratio (1:72) with net return of Rs. 45, 927/= as against the cropping pattern Jute-Paddy-Rajma (Rs. 25, 491 /=), Jute-Paddy-Mustard (Rs. 19, 896 /=) and JutePaddy-Pea (Rs. 17, 033 /=) during the year 2002 - 2003. Same trend of beneficial effect was found in previous year also. Farmers are very much convinced with the result and motivated to the designated crop sequence and the jute varieties Development in USA Researches are going on in the USA on different aspects of kenaf production e.g. variety selection, the yield, resistance to diseases, nematodes, pests and weed control. In the U.S., the varieties used most extensively are those developed by ARS researchers in Florida - 'Everglades 41' and 'Everglades 71'. Both varieties are resistant to anthracnose. In the southern U.S., kenaf emerges and grows so rapidly that it competes effectively with weeds. Growers and researchers of the U.S. have found kenaf to be resistant to most plant diseases. The variety development work done by the ARS in Florida was very effective in breeding and selecting for anthracnose resistance. There has been little economic damage to kenaf by insects in experimental production fields in U.S.A.

Interspecific hybridisation and its limitations Strains resistant to jute leaf mosaic disease have been developed from interspecific crosses between C. trilocularis (wild species) and C. capsularis. These strains are now in the pipe line. Improvements to cultivated jute through varietal development and intraspecific hybridisation have been under way since the beginning of this century by multidirectional crossing, followed by selection, amongst the more promising types (Ghosh,1983). However, these approaches have some limitations for introducing specific genes for specific purposes. On the other hand, C. capsularis and C. olitorius, the two cultivated species, possess the characteristics in their own right yet clearly need to be hybridised. Several attempts using terms of sexual hybridisation between these two species have been made (Table 1). A number of researchers reported hybridisation between the two species C. capsularis and C. olitorius to be unsuccessful (Finlow, 1917, 1921, 1923; 1924). Banerjee and Datta, 1960; Patel and Datta, 1960; Datta et al. 1960). Many however (Islam and Rashid,1961; Swaminathan et al. 1961; Choudhuri and Mia, 1962; Mia and Shaikh,1967) have claimed to have succeeded in producing hybrids. However, all those putative hybrids showed dominance of the female parent in F1 and F2 generations. Raut and Naik (1983) claimed to have followed the hybrid progenies up to F3 generation but there was no further report of exploitation of those hybrid at later generation. This is probably because of the fact that in later generation the entire population resembled the female parent. The F1 and the following hybrid generations (Table 1) was possible to obtain only when C. olitorius was used as a pistillate parent. It has not been possible so far to obtained a hybrid using C. capsularis as a female parent although only one successful fertilization was reported by Bhaduri and Bairagi (1968) in this direction. Whether or not the cytoplasmic factors of C. capsularis and C. olitorius have any role in inhibiting the fruit set in case of C. capsularis (used as female parent) or induction of apomixis or chromosome elimination for C. olitorius (as the female parent) yet to be determined. Table 1 A summary of interspecific hybridisation attempts (conventional methods) between C. olitorius and C. capsularis. Crossing details Normal crossing Normal crossing Results obtained No fruit setting No fruit setting Reference Finlow (1917,1921,1923 Howard (1924) Bhaduri and Chakravarty (1948) Srinath and Kundu(1952) Ganesen et al. (1957) Banerjee and Datta, 1960 Patel and Datta (1960) Islam and Rashid (1961)

Implanting the style of C. F1 showed maternal inheritance capsularis on the style of only C. olitorius Normal crossing Premature cessation of embryo growth Normal crossing Hybrid embryo failed to germinate Normal crossing No hybrid Wide range of cultivated and A few non-viable seeds obtained wild types used Smearing the pedicel of the F2 and F3 showed female parent flower with IAA traits Receprocal grafting of the parental species and use of Fruit could not attain maturity X-ray irradiated pollen for hybridisation

Swaminathan et al.(1961)

Crossing between autotetra- Fruit could not attain maturity ploid cultivated and wild jute Normal crossing F2 showed mostly female parent traits Normal crossing F1 maternal in appearance Normal crossings, both of F1 maternal and F2 data not given diploid and tetraploid jute Growth hormones and F1 and F2 hybrid showed maternal embryo culture inheritance only Application of IAA at the F1 similar to female parent ; no pedicel of the pollinated segregation in F 2 flowers Using gamma- irradiated F1 maternal and F2 data was not pollen given Normal crossing F1 and F2 maternal in appearance A wide range of wild species F1 maternal and no segregation in F2 and cultivars used Normal crossing Claimed success but no variety ever released Reciprocal grafting of the parental species; application F2 mostly maternal in appearance of IAA embryo culture

Datta et al. (1960) Swaminathan and Iyer (1961) Iyer et al. (1961) Choudhuri and Mia (1962) Islam (1964)

Frost and Bose (1966) Mia and Shaikh (1967) Bhaduri and Bairagi ( 1968) Arangzeb and Khatun (1980b,1981,1982a,1983 Raut and Naik (1983) Hoque et al. (1988)

Improvement of jute through conventional sexual hybridization between two cultivated species, Corchorus capsularis and C. olitorius could not be achieved due to the absence of intermediate characters in F1 hybrids. Whilst these two species can be crossed sexually, F1 and F2 plants contained only maternal characters. This was confirmed by isoenzyme profiles. Where F 1 plants showed bands similar to C. olitorius ( ). Unique hybrid bands or those of male parent C. capsularis ( ) were missing. It was suspected that failure to obtain true hybrids was either due to apomixis or chromosome elimination. Biotechnological approaches for crop improvement In the field of biotechnology, some developments have been made on jute, kenaf and mesta. The culture of plant cells, tissues and organs under defined laboratory conditions has both pure and applied applications. A unique feature of plant cells is that they exhibit totipotency. This enables intact, fertile plants to be regenerated from tissues which are given the appropriate stimuli from exogenously supplied growth regulators, such as auxins and cytokinins, in the culture medium. The ability to regenerate plants is fundamental to the multiplication of elite individuals by micropropagation. Additionally, the genetic engineering of plants, through exposure of somaclonal variation, somatic hybridization by protoplast fusion and transformation involving Agrobacteriummediated gene delivery or direct DNA uptake, also necessitates reproducible plant regeneration. Tissue culture For this purpose protocols for jute explant and protoplast culture/regeneration systems were developed. Plant regeneration was unachievable from cotyledons (without petioles), hypocotyls and root explants. However, callus could readily be induced from these explants using various combinations of growth regulators in MS-based media. This eventually gave rise to rhizogenesis. Somatic embryos were produced from explants of C. capsularis in the presence of 2,4-D and BAP but shoot regeneration proved impossible. Multiple shoots could be regenerated from internodes,

shoot apices, zygotic embryos, leaf petioles and cotyledons with attached petioles. In these cases, plant regeneration occurred via embryogenesis which was confirmed by histological examination. The effects of non-ionic surfactants (Pluronic F-68, Triton X-100 or Tween 20), on shoot regeneration for jute (C. capsularis) cotyledons with attached petioles and growth of transformed roots was assessed. Supplementation of culture media with 0.001-0.5% (w/v) Pluronic F-68 increased the mean percentage of cotyledons producing shoots and the mean number of shoots/cotyledons. By contrast, Tween 20 produced maximal effects at 0.001% (v/w) with inhibition of shoot formation at 0.5% (v/w). In both cases, phenotypically normal plants were recorded which could be grown to maturity. Culture of cotyledons with 0.001% (v/w) Triton X-100 similarity increased both the percentage of cotyledons producing shoots and the number of shoots/cotyledons. However, these shoots did not develop into mature plants. The effects of these non-ionic surfactants, on the jute transformed roots responded similarly for root growth. These results were linked to the hydrophilic-hydrophobic (HLB) balance of individual surfactants, which, in turn, determined their cell permeabilising properties and consequently, their putative effects on morphogenesis. Plant regeneration was also established from the cotyledons (with attached petioles) of kenaf and mesta. Regenerated plants were successfully transferred into soil. These established systems are now ready for gene transfer.

Somatic hybridization As interspecific hybridisation between two cultivated species of jute was unsuccessful, somatic hybridisation was attempted. Somatic hybridisation was attempted between C. capsularis and C. olitorius as sexual hybridization between two cultivated species could not be achieved. For this purpose, protoplasts isolated from cotyledons, hypocotyls and cell suspensions of C. capsularis and C. olitorius and callus production and somatic embryogenesis was induced from protoplasts. A high frequency heterokaryon formation between C. capsularis and C. olitorius was achieved via electrofusion. These heterokaryons, endured sustained division to form colonies and calli. Multiple shoots regenerated from various explants of C. capsularis and C. olitorius as it is a prerequisite of transformation. Protoplast isolation and culture Protoplasts were isolated from cotyledons, hypocotyles and cell suspensions of C. capsularis and C. olitorius. Mucilage-related difficulties in protoplast isolation were overcome by controlling the growth conditions of the seedlings prior to isolation. Seedlings required continuous intense illumination and a high temperature (28C). Cotyledon protoplasts of both of these species were divided in K8P medium with agarose bead culture, whereas, cell suspension protoplasts were divided in both liquid and agarose media. Necrosis development was noticed in the dividing protoplasts of C. olitorius. The presence of an enriched oxygen atmosphere inhibited necrosis of the developing colonies and thereby increased plating efficiency for jute protoplasts. Callus could be readily initiated from the protoplasts of cotyledons, hypocotyls and cell suspensions of C. capsularis and C. olitorius. The cotyledon and hypocotyl protoplast-derived callus of both species became compact and nodular and exhibited rhizogenesis. Somatic embryogenesis was also induced from cotyledon protoplast-derived callus of C. capsularis. Somatic fusion A high frequency heterokaryon formation between C. capsularis and C. olitorius was achieved via electrofusion. These heterokaryons, endured sustained division to form colonies and calli. A selection system for somatic hybrids was established based on differential growth tolerances (division) of cotyledon and cell suspension protoplasts in liquid media. Cell suspension clones of

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the (putative) hybrid showed green pigmentation and vigorous growth which was absent for the parents. In isoenzyme profiles (acid phophatase), hybrid cell lines showed prominent hybrid bands and included both parental bands. Genetic transformation Transgenic hairy roots were induced in jute cotyledons and hypocotyls of both C. capsularis and C. olitorius using Agrobacterium rhizogenes strains and somatic embryos were obtained from these tissue. A direct transgenic plant regeneration protocol was developed from A. tumefaciens infected explants. A protocol for plant regeneration from kenaf and mesta have been developed using cotyledon explants. Transgenic roots were induced in cotyledons and hypocotyls of both C. capsularis and C. olitorius using wild and engineered (antibiotic resistant) strains of Agrobacterium rhizogenes. These roots were confirmed as transformed with positive opines and NPTII assays. Additionally, the engineered strains (R1601 and LBA 9402 pBin 19) showed resistance to 100 g/ml of kanamycin sulphate whwereas the non-transformed seedling roots could only survive upto 25 g/ml of kanamycin. Somatic embryos were obtained from these tissues in the presence of 2,4-D, but did not regenerate into plants after transfer to 2,4-D free medium. A direct transgenic plant regeneration protocol was established for A. rhizonenes-infected cotyledons stimulated by the presence of non-ionic surfactants. These plants showed abnormal morphology including hairy root production from the leaves and stems in the presence of the surfactant (Pluronic F-68). Although these plants did not show a positive reaction to opine and NPTII assays, the presence of adventitious roots on leaves and stems strongly suggested a transformed nature. An efficient and reproducible protocol for transformation of kenaf varieties has been developed, which showed the integration of marker genes. DNA finger printing DNA fingerprinting of 18 jute genotypes composed of popular varieties and some promising lines from both C. olitorius and C. capsularis have been done using RAPD and SSR techniques. A molecular marker for low temperature tolerance has developed and with this marker it was possible to identify a gene. A non-radioactive differential display was established for low temperature stress tolerance in jute. RT-PCR was conducted using the total mRNA isolated from three jute strains differing in the low-temperature tolerance and the gene expression was possible to determined. Conclusion Several high yielding varieties of jute, kenaf and mesta were developed using traditional methods. However, these techniques have some limitations. For example, sexual hybrid was not obtained between two cultivated species of jute using conventional methods which could be overcome through somatic hybridization. Development of varieties with some specific genes for example, Bt genes are not yet introduced in jute, kenaf or mesta. Biotechnological approaches are therefore needed for developing varieties with the insertion of specific genes beside the traditional approaches. References Arangzeb, S. and Khatun, A. (1980a) A short note on interspecific hybrids between C. trilocularis L. and C. capsularis L. Bangladesh. J. Jute Fib. Res. 5, 85-89. Arangzeb, S. and Khatun, A. (1980b) Interspecific crosses of vegetable types and other closely related C. capsularis with standard wild C. olitorius varieties by applying various

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techniques. Ann. Rep. Bangladesh Jute Research Institute (Agric.), Dhaka, Bangladesh. pp. 49-50. Arangzeb, S. and Khatun, A. (1981) Ibid, pp. 55-56. Arangzeb, S. and Khatun, A. (1982a) Ibid, p. 65. Arangzeb, S. and Khatun, A. (1982b) Interspecific hybridisation and role of apomixis in the two species of Corchorus. 6th and 7th Annual Bangladesh Science Conference, Dhaka, Bhaduri, P. N. and Bairagi, P. (1968) Interspecific hybridisation in jute (Corchorus capsularis x C. olitorius) Sci. Cult. (Calcutta) 34, 355-357. Datta, R. M., Dana, S. K. and Banerjee, S. N. (1960) Investigation on the interspecific hybridization between autotetraploids of the cultivated jute species (Corchorus olitorius L. and C. capsularis L. ) and the failure of viable seed formation in them. Genet. Iber. 12, 1-32. F. B. d'Utra Vaz, Slamet, I. H., Khatun, A ., Cocking, E. C. and Power, J. B. (1992) Protoplast culture in high molecular oxygen atmospheres. Plant Cell Reports. 11 : 416-418. Finlow, R. S. (1917) Historical notes on experiments with jute in Bengal. Agric. J. India, 12, 3-29. Finlow, R. S. (1921) Historical notes on experiments with jute in Bengal. Agric. J. India, 16, 265279. Finlow, R. S. (1923) Note on the work on fibre selection in Bengal. Agric. J. India, 3, 138. Frost, S. and Bose, S. (1966) An investigation of the phenolic compounds in two species of jute (Corchorus olitorius and Corchorus capsularis) and their supposed hybrids, using the thin layer chromatographic technique. Hereditas, 55, 183-187. Ganesen, A. T., Shah, S. S. and Swaminathan, M. S. (1957) Cause for the failure of seed-setting in the cross Corchorus olitorius x C. capsularis. Curr. Sci. 26, 292-293. Howard, A. (1924) Crop Production in India, Oxford University Press. pp. 138. Islam, A. S. (1964) A rare hybrid combination through application of hormone and embryo culture. Nature, 210, 320. Islam, A. S. and Rashid, A. (1961) First successful hybrid between the two jute yielding species, C. olitorius x C. capsularis. Nature, 185, 258-260 Iyer, R. D., Sulbha, K. and Swaminathan, M. S. (1961) Fertilization and seed development in crosses between Corchorus olitorius and C. capsularis Indian J. Gen. Plant Breed. 21, 191200.

Khatun A, M.R. Davey, J.B. Power and K.C. Lowe, 1993. Stimulation of shoot regeneration from jute cotyledons cultured with non-ionic surfactants and relationship to physicochemical properties. Plant Cell Rep., 13: 49-53. Khatun, A., 1993. The Genetic Manipulation of Jute (Corchorus) Species. Ph.D. Thesis. Dept. of Life Sci. University of Nottingham, UK, pp: 44-81.

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Khatun, A., S. Mahboob and M. S. Alam, 2001-2002. Attempts to obtain plant regeneration system from the C. olitorius. Annual Report. Bangla. Jute Res. Inst., Manik Mia avenue, Dhaka, Bangladesh. pp: 60-63. Khatun, A. and Z. Naher, 2001-2002. Optimization of culture conditions for plant regeneraton from kenaf explants of kenaf and mesta. Annual Report. Bangla. Jute Res. Inst., Manik Mia avenue, Dhaka, Bangladesh, pp 57-58.
Khatun, A ., Laouar, L., Davey, M. R., Power, J. B. Mulligan B. J. and Lowe, K. C. (1992) Pluronic F-68 enhances growth and morphogenesis of cultured tissues. Abstract 53. Abstract posters proceedings, Lancaster Meeting on Physiology, growth and Development of Plants and cells in culture. Lancaster. England. Khatun, A. Davey, M. R., Power, J. B. Cocking, E. C. (1990) Transformation of jute (Corchorus spp.) using Agrobacterium rhizogenes conferring a chimaeric neomycin phosphotransferase (NPTII) gene. Abstract A2-82. Proceedings of the International Congress on Plant Tissue and Cell Culture, Amsterdam, The Netherlands. Mia, M. M. and Shaikh, A. Q. (1967) Gamma radiation and interspecific hybridisation in jute (Corchorus capsularis L. and C. olitorius L.) Euphytica, 16, 61-68. Srinath, K. V. and Kundu, B. C. (1952) Cytological studies of pollen-tube growth in reciprocal crosses between Corchorus capsularis L. and C. olitorius L. Cytologia, 17, 219-223. Swaminathan, M. S. and Iyer, R. D. (1961) Skewed recombination in a rare interspecific jute hybrid. Nature, 192, 893-894. Swaminathan, M. S., Iyer, R. D. and Sulbha, K. (1961) Morphology, cytology and breeding behaviour of hybrids between Corchorus olitorius and C. capsularis. Curr. Sci. 30, 67-68.

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