Interspecific hybridization of Trifolium alexandrinum with T . constantinopolitanum using embryo rescue

Roy, A. K.; Malaviya, D. R.; Kaushal, Pankaj; Kumar, Brijesh; Tiwari, Aparna

doi:10.1007/s00299-004-0759-1

Cited by 26 publications

(29 citation statements)

References 14 publications

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“…The regenerated plantlets in the culture tubes were kept out of the culture room for 2-3 d at room temperature (30°C). The plants were taken out of the culture tubes, freed of media and kept for an additional day with the roots submerged in sterilized distilled water containing crushed nodules from T. alexandrinum plants growing in field following Roy et al (2004Roy et al ( , 2005 and covered with cellophane paper to maintain high humidity. These plants were protected from direct sunlight for the first 3-5 d in the field.…”

Section: Methodsmentioning

confidence: 99%

In vitro Callusing and Regeneration in Trifolium resupinatum-A Fodder Legume

et al. 2006

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SummaryIn vitro regeneration protocol was developed in Trifolium resupinatum, an important fodder legume in Northwestern India. Out of 6 explants (leaf, petiole, hypocotyl, cotyledon, collar and root) tried for callus induction in 2 different media, good response was observed from hypocotyl and root explants in 'A' medium. Successful regeneration was obtained from callus induced from hypocotyl, cotyledon and root in 'A' followed with subculture in 'E' medium. One of the regenerated plantlet was characterized and compared with mother plant for isozyme-banding pattern. Wide variation for isozyme banding pattern for SOD, Peroxidase and GOT enzyme indicates the regenerant to be a somaclonal variant. Thus, in vitro regeneration will help in widening the genetic base of the species.Key words Trifolium resupinatum, Fodder, Regeneration, Tissue culture.Plant tissue, cell and protoplast cultures are useful tools for crop improvement, especially as supplementary means of inducing variability through tissue culture instability and somaclonal variations (Scowcroft 1984). Trifolium resupinatum (Shaftal or Persian clover) is an important cultivated winter annual fodder legume grown widely in subtropical and sub temperate areas of India. The species has previously been frequently introduced in different parts of Europe and cultivated as fodder crop in northwestern subtropical zones. It is a multicut crop and is favoured by farmers for its high quality, good yield and tolerance to biotic and abiotic stress.Narrow genetic base in this crop hampers its genetic improvement programme. The conventional breeding efforts have failed to generate variability for making effective selection. In this study, efforts were made to develop and characterize somaclonal variants in this crop by using in vitro tissue culture methods. Materials and methodsHealthy seeds of T. resupinatum (SH 97-49) were surface sterilized using 0.1% HgCl 2 for 2 min followed by 4-5 rinsing with sterile water and germinated on basal MS medium (Murashige and Skoog 1962) containing 3% sucrose and 0.7% agar without any growth regulators. Six explants viz., leaf, petiole, hypocotyl, cotyledon, collar and root were taken from 21-23 d old seedlings. Two combinations of auxin and cytokinin in the medium were studied for callus induction efficiency. The medium 'A' contained L2 basal medium (Phillips and Collins 1984) supplemented with 0.05 mg/l NAA and 0.1 mg/l BAP, whereas medium 'A-1' contained L2 basal medium supplemented with 0.1 mg/l BAP. Initially the explants were kept in dark for callus induction and after 5 d of inoculation the cultures were provided with a photoperiod of 10 h. After shoot emergence, the light

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Section: Methodsmentioning

confidence: 99%

In vitro Callusing and Regeneration in Trifolium resupinatum-A Fodder Legume

et al. 2006

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“…Hybridisation with T. apertum (2n = 16) via in vitro embryo rescue generated 20 hybrid plants showing introgression of various desirable traits, including late flowering [96]. With the aid of embryo rescue, hybridisation was also successfully achieved with T. constantinopolitanum (2n = 16) [97] and T. resupinatum [98]. Some of the hybrid plants from the latter cross showed late flowering and the higher survival rate in the field compared to T. alexandrinum.…”

Section: T Subterraneum and T Alexandrinummentioning

confidence: 89%

Development of Genomic Resources in the Species of Trifolium L. and Its Application in Forage Legume Breeding

2012

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Clovers (genus Trifolium) are a large and widespread genus of legumes. A number of clovers are of agricultural importance as forage crops in grassland agriculture, particularly temperate areas. White clover (Trifolium repens L.) is used in grazed pasture and red clover (T. pratense L.) is widely cut and conserved as a winter feed. For the diploid red clover, genetic and genomic tools and resources have developed rapidly over the last five years including genetic and physical maps, BAC (bacterial artificial chromosome) end sequence and transcriptome sequence information. This has paved the way for the use of genome wide selection and high throughput phenotyping in germplasm development. For the allotetraploid white clover progress has been slower although marker assisted selection is in use and relatively robust genetic maps and QTL (quantitative trait locus) information now exist. For both species the sequencing of the model legume Medicago truncatula gene space is an important development to aid genomic, biological and evolutionary studies. The first genetic maps of another species, subterranean clover (Trifolium subterraneum L.) have also been published and its comparative genomics with red clover and M. truncatula conducted. Next generation sequencing brings the potential to revolutionize clover genomics, but international consortia and effective use of germplasm, novel population structures and phenomics will be required to carry out effective translation into breeding. Another avenue for clover genomic and genetic improvement is interspecific hybridization. This approach has considerable potential with regard to crop improvement but also opens windows of opportunity for studies of biological and evolutionary processes.

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“…For Rhizobium inoculation, nodules collected from the old plants from the field were macerated in distilled water and the root of regenerated plants was dipped for overnight following Roy et al (2004). The plants were then transferred to pots having soil and sand in equal proportion.…”

Section: ⎯⎯⎯⎯mentioning

confidence: 99%

In vitro regeneration of Trifolium glomeratum

Kaushal¹,

Tiwari²,

Roy³

et al. 2006

Biol. plant.

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In vitro regeneration of Trifolium glomeratum, a leguminous forage species, was attempted through leaf, petiole, cotyledon, hypocotyl, collar and root explants and two media combinations. Root and collar explants showed no callus induction. Medium with 0.05 mg dm -3 α-naphthaleneacetic acid (NAA) and 0.10 mg dm -3 N 6 -benzyladenine (BA) was more effective for hypocotyl explant whereas cotyledon and petiole explant were more responsive to 5.0 mg dm -3 NAA and 1.0 mg dm -3 BA. Friable, green calli obtained from petiole explant on this medium showed organogenetic potential. Modified root-inducing medium having 0.21 mg dm -3 indole-3-acetic acid and 2.5 % sucrose was successful for root induction and plantlets were successfully transferred to field after hardening and Rhizobium inoculation.Additional key words: auxins, clovers, cytokinins, tissue culture. ⎯⎯⎯⎯The genus Trifolium, comprises of about 300 species of which twenty-five are important as cultivated forages and pasture crops for both wild and domestic animals. T. glomeratum, an annual diploid species (2n=2x=16) with good forage quality, is frequently inhabited in dry places of Europe. The species possesses close affinity with agriculturally important species (e.g. T. repens, T. alexandrinum) (Malaviya et al. 2005), and hence, may be utilized as donor for desirable traits such as resistance to root rot and stem rot (Bhaskar et al. 2002). Efficient in vitro regeneration protocol is a prerequisite to conduct biotechnological experiments (Espino et al. 2004, Zapata et al. 2004. Hence, the present investigation was carried out to study the effect of different media (growth regulator combinations) and explant towards in vitro response and to develop a suitable in vitro regeneration protocol.Healthy seeds of Trifolium glomeratum L. (accession EC 401700) were surface sterilized using 0.1 % HgCl 2 for 1 min followed by two to three washings in distilled sterilized water. Seeds were germinated on MS 0 medium (Murashige and Skoog 1962), devoid of any growth regulators, containing 3 % sucrose and agar (0.7 %). For callus induction, shoot induction and somatic embryogenesis, L2 basal composition as suggested by Phillips and Collins (1984) was used. Callus induction was attempted on 'A' medium [having 0.05 mg dm -3 α-naphthaleneacetic acid (NAA), and 0.10 mg dm -3 N 6 -benzyladenine (BA)] as well as 'D' medium (having 5.0 mg dm -3 NAA and 1.0 mg dm -3 BA). In shoot inducing medium 'E', NAA and BA were 0.0008 mg dm -3 and 0.15 mg dm -3 respectively. For somatic embryogenesis, 2,4-dichlorophenoxyacetic acid (2,4-D; 0.001 mg dm -3 ) and adenine (3.225 mg dm -3 ) were used in somatic embryogenesis inducing medium (SEIM) medium. Root induction was done on RL1 medium consisting of RL basal medium (Phillips and Collins 1984) and 0.21 mg dm -3 indole-3-acetic acid (IAA) and 2.5 % sucrose. Six explants viz., leaf, petiole, hypocotyl, cotyledon, collar and root were taken from 20 to 30-d-old healthy seedlings and inoculated in callus inducing media. The cultures were maintained...

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Interspecific hybridization of Trifolium alexandrinum with T . constantinopolitanum using embryo rescue

Cited by 26 publications

References 14 publications

In vitro Callusing and Regeneration in Trifolium resupinatum-A Fodder Legume

In vitro Callusing and Regeneration in Trifolium resupinatum-A Fodder Legume

Development of Genomic Resources in the Species of Trifolium L. and Its Application in Forage Legume Breeding

In vitro regeneration of Trifolium glomeratum

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