P.N. Krishnan

Jawaharlal Nehru Tropical Botanic Garden and Research Institute, Tiruvananantapuram, Kerala, India

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Publications (26)15.65 Total impact

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    ABSTRACT: While using the protocols for the long-term conservation, assessment of true-to-type regenerants from cryopreserved materials is important. In the present study, no significant variation in banding pattern was noticed in the RAPD profile of cryopreserved and control samples of shoot tip-derived Kaempferia galanga plants. But in the case of somatic embryoderived samples, some variation was observed in the banding pattern (16.2% polymorphism), though the plants were phenotypically similar. Recovery of cryopreserved somatic embryos in K. galanga was preceded through a callus phase, from which secondary embryos were induced later. The RAPD profile of somatic embryo-derived samples revealed that variation was due to the occurrence of callus phase. As minor genetic variations arising in in vitro cultures without marked phenotypic changes are considered to be beneficial for diversity conservation and sustainable utilization, cryopreserved somatic embryos of K. galanga would serve as an alternative for generating and maintaining genetic diversity of this medicinal wealth.
    No preview · Article · Jan 2015

  • No preview · Conference Paper · Dec 2014
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    ABSTRACT: A rapid in vitro propagation protocol for large-scale cultivation of Lagerstroemia speciosa L., an important medicinal and ornamental tree was established using nodal explants in Schenk and Hildebrandt (SH) medium optimized with various concentration and combinations of auxins and cytokinins. The study revealed, for the first time, accumulation of corosolic acid (CRA), an anti-diabetic and anti-obese pentacyclic triterpene in the in vitro plantlets. Multiple shoot induction was maximum in SH medium supplemented with 4.44 µM Benzyl Amino Purine (BAP) and shoot initials, when transferred to SH medium supplemented with 2.22 µM BAP, resulted in shoot multiplication and elongation. Rooted plantlets, after 2 weeks of hardening in mist-house and 3 months of rearing under shade-net, transferred to forest segments showed successful establishment (100 %). In a short duration of 24 weeks, ~2,450 hardened plantlets were obtained from a single nodal explant. The ISSR banding profiles of the regenerants and the mother plant were highly monomorphic and similarity matrix (0.96–0.99) confirmed genetic fidelity of the clones. Chemical analysis using HPLC showed that CRA content is more in mature (0.012–0.062 % DW) than young leaves (0.004–0.007 % DW) of the in vitro derived plantlets. The differential synthesis of CRA in micropropagated and mother plant was studied using gene expression profiles. The expression of upstream rate limiting genes in terpenoid biosynthesis was relatively high in young leaves while downstream Cytochrome P450 hydroxylase (CYP450H), catalyzing the final step(s) in CRA synthesis, was more in mature leaf, suggesting a strong correlation between observed CRA variation and gene expression. The study signifies the role of in vitro methodology for true-to-type regeneration and its possible utility for biosynthesis of CRA, throughout the year.
    No preview · Article · Dec 2014 · Plant Cell Tissue and Organ Culture
  • T.S Preetha · A.S. Hemanthakumar · P.Padmesh · P.N.Krishnan
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    ABSTRACT: An efficient in vitro propagation protocol was standardized in K. galanga, wherein shoot cultures were raised from rhizome with axillary bud explants in Murashige and Skoog (MS) medium supplemented with different hormonal regimes. Maximum 10.6±0.83 multiple shoots per explants were obtained in MS medium supplemented with 4.0 mgl-1 6-benzyl adenine (BA) along with 1.0 mgl-1 each of α-naphthaleneacetic acid (NAA) and kinetin. Subsequent 2-3 subculture passages enhanced the shoot multiplication rate to 3-4 times. The multiple shoots thus produced were having individual root system and elongated substantially when kept in the same medium for 4-6 weeks. There was no need of elongation and root induction phase and this two-step multiplication procedure reduced the period of plantlet production compared to the earlier protocols. The regenerated plants after a short hardening phase got established in the field at 80-90% efficiency and their genetic uniformity was confirmed through ISSR analysis. The protocol thus standardized is an efficient method for rapid propagation of this high value medicinal species by which at least 30 shoots per explants can be produced within 12 weeks duration. It offers the possibility of raising physiologically uniform plants for easy dissection of shoot tip meristems which can be efficiently utilized for subsequent long-term conservation through cryopreservation.
    No preview · Article · Dec 2014

  • No preview · Conference Paper · Jan 2014

  • No preview · Conference Paper · Dec 2013
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    ABSTRACT: An efficient protocol has been devised for the propagation and field establishment of Eulophia cullenii (Wight) Bl., a terrestrial orchid having ornamental potentialities, and is critically endangered in Western Ghats, India. Seeds extracted from 60–90-d-old capsules germinated in ½ MS, ¼ MS, Knudson C, or Mitra liquid medium developed into 1.4–2.5-mm-diameter protocorms in 60 d. Supplementation of organic additives like coconut water, peptone, yeast extract, and casein acid hydrolysate (CH) significantly enhanced protocorm growth. Upon subculture onto agar-gelled Mitra medium fortified with 0.05% CH, 56% of protocorms regenerated into shoots through the formation of linear mini-rhizomes. The regenerated shoots grew vigorously in ½ MS, producing new rhizomes. Mature rhizomes from axenic seedlings produced maximum (13 ± 1.4) shoots/whole rhizome in ½ MS fortified with 44.4 μM 6-benzylaminopurine (BAP), in 120–150 d. Horizontal and longitudinal halves of the rhizome also gave multiple shoots (6–8.5) in the presence of 44.4 μM BAP. Shoots or shoot clumps sub-cultured onto ½ MS basal medium produced roots followed by rhizomes in 60–150 d. Seedlings with mature rhizomes showed 70% establishment in the nursery and added a new rhizome at the end of one growth cycle. An average of 70.6% of the rhizomes originating from seedlings during the second growth cycle sprouted to produce new shoots, when planted in the native localities. Asymbiotic germination and cloning through rhizomes thus can provide a large number of vigorous plants of E. cullenii for ornamental exploitation as well as eco-restoration, if rhizome as storage organ is ensured in the propagule.
    No preview · Article · Nov 2013 · In Vitro Cellular & Developmental Biology - Plant

  • No preview · Conference Paper · Oct 2013
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    ABSTRACT: Musa acuminata Colla, the wild progenitor of banana, has a long evolutionary history intervened by human activity. Forty three accessions of M. acuminata ssp. burmannica, collected from 3 wildlife sanctuaries of Agastyamalai Biosphere Reserve of Western Ghats, India were studied for their genetic diversity and differentiation using random amplified polymorphic DNA (RAPD) markers. The results indicated a relatively high level of genetic diversity in Musa accessions at the species [Nei's gene diversity (H)=0.47; Shannon information index (I)=0.66)] as well as population level (H=0.42; I=0.61). A relatively high degree of genetic differentiation was also observed among the populations (GST=0.30). Moreover, analysis of molecular variance (AMOVA) showed the existence of vast (81%) genetic variation within the populations. The Mantel test revealed a significant correlation (Rxy=0.54; P<0.001) between the geographic distance and the genetic distance of these populations. The genetic variability among these accessions suggests that the management for the conservation of the genetic diversity in M. acuminata should aim at preserving each and every accession at Agastyamalai Biosphere Reserve.
    No preview · Article · Oct 2013 · Indian Journal of Biotechnology
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    A. S. Hemanthakumar · T. S. Preetha · P. N. Krishnan · S. Seeni
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    ABSTRACT: Zygotic embryos excised from immature green fruits of the rattan palm, Calamus thwaitesii and cultured for 16 weeks under optimum culture conditions in Murashige and Skoog (MS) medium supplemented with 31.67 μM 2,4-dichlorophenoxyacetic acid (2,4-D) and 35.23 μM 2,4,5-trichlorophenoxyacetic acid (2,4,5-T) produced mixed (compact and friable) calli at 70 and 92 % rates. The semi-friable part of the callus (~500 mg) separated and subcultured in medium containing 2.22 μM 6-benzyladenine and 1.07 μM α-naphthalene acetic acid produced groups of 10.37 ± 0.60–21.52 ± 0.48 discrete globular embryoids of varied size in 6–8 weeks. Calli raised in presence of 2,4,5-T were relatively more prolific, friable and embryogenic than those induced by 2,4-D. Embryoids (2.0–3.0 mm) isolated and cultured in basal medium germinated into plantlets at 65 % efficiency while the immature (0.5–2.0 mm) ones produced calloid structures. Approximately 15 % of the in vitro plantlets raised from the 2,4-D-induced embryogenic calli produced secondary immature embryoids on the sheath and lamina parts of leaves which were isolated and cultured in basal medium developed into rooted plantlets at 62 % rate in 12–16 weeks. The continued growth of the embryo-derived callus through successive subcultures together with differentiation of embryoids into plantlets, and the formation of immature embryoids on in vitro plantlets in MS basal nutrient medium reports for the first time a reliable method of producing at least 116 plants from a single embryo in a year. Rooted plantlets treated with 50 % glycerin survived at 78 % rate after hardening and 82.7 % of the hardened plants reintroduced into forest segments showed uniform growth free of morphological abnormalities after 3 years of observation. In addition to embryogenesis, cryopreservation of the zygotic embryos through simple drying and encapsulation–dehydration methods resulting 60–70 % recovery rates also offers another option for long-term conservation and sustainable utilization of this plant genetic resource.
    Full-text · Article · Jun 2012
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    ABSTRACT: Musa acuminata ssp. burmannica, one of the wild progenitors contributing ‘A genome’ to the present-day dessert bananas, has a long evolutionary history intervened by human activities. In this study, ISSR markers were used to analyze the pattern of genetic variation and differentiation in 32 individuals along with two reference samples (viz., Musa acuminata ssp. burmannicoides, var. Calcutta 4 and Musa balbisiana) of wild Musa, which corresponded to three populations across the biodiversity-rich hot spot of southern Western Ghats of India. High levels of genetic diversity were revealed both at the species and population levels, using Nei’s diversity indices. The hierarchical analysis of molecular variance showed pronounced genetic differentiation, as 96 % of the total variance was fixed within population and only 4 % among populations. Nei’s genetic differentiation coefficient (G ST = 0.1823) and low gene flow (Nm = 1.18) further confirmed this. The positive correlation (Mantel test) between geographic distance and genetic distance (r = 0.338 P < 0.001) indicates geographic isolation as one of the key factors in shaping the population genetic structure. Grouping of individuals was largely in conformity with their spatial distribution, which was confirmed by UPGMA cluster analysis and PCA scatter plot clustering all 32 individuals into three major groups along a geographical gradient. The discontinuous distribution and dwindling population due to habitat fragmentation are serious threats to prevailing genetic diversity in this species. Conservation measures based on diversity pattern are suggested for long-term preservation and sustainable utilization of this precious genetic resource. Key message A diverse germplasm of Musa acuminata ssp. burmannica exists in southern Western Ghats as a possible repository of useful resistant traits, which can be effectively utilized for crop improvement.
    Full-text · Article · May 2012 · Plant Cell Reports
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    R.K. Radha · William S. Decruse · P.N. Krishnan

    Preview · Chapter · Mar 2012

  • No preview · Conference Paper · Mar 2012

  • No preview · Conference Paper · Jan 2012
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    ABSTRACT: In vitro propagation of diploid land race of Musa cv. ‘Ambalakadali’ was achieved through organogenesis both from shoot primordial meristems and male floral apices taken out from mature disease free field grown plants. Healthy cultures were successfully initiated from shoot primordial meristems when independent dosage of BAP at 4.0-mg/l was given in MS nutrient media. The inflorescence apices responded the most to the in vitro culture conditions at 6.0-mg l-1 BAP in MS media itself. In both the cases, culture proliferation requires comparatively lower dosages of BAP. Subsequent cultures displayed enhancement in rate of shoot initiation and multiplication in the inflorescence derived cultures. Transfer of shoots (4-5 cm) into MS basal medium containing 0.1% activated charcoal (AC) favoured generation of healthy shootlets and formation of profuse roots during a period of 4 weeks. The plantlets (15 cm) after acclimatization in green house were distributed to local farmers for field evaluation. Plants flowered normally within 12 months and produced uniform characters as that of the mother plants.
    Full-text · Article · Mar 2011
  • Peringattulli Narayanan Krishnan · S. W. Decruse · R. K. Radha
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    ABSTRACT: Climate change, alien species, and use of land for intensive farming and development are causing severe threat to the plant genetic diversity worldwide. Hence, conservation of biodiversity is considered fundamental and also provides the livelihoods to millions of people worldwide. Medicinal plants play a key role in the treatment of a number of diseases, and they are only the source of medicine for majority of people in the developing world. The tropical regions of the world supply the bulk of current global demand for “natural medicine,” albeit with increasing threat to populations in the world and its genetic diversity. India is a major center of origin and diversity of crop and medicinal plants. India poses out 20,000 species of higher plants, one third of it being endemic and 500 species are categorized to have medicinal value. The Western Ghats is one of the major repositories of medicinal plants. It harbors around 4,000 species of higher plants of which 450 species are threatened. Currently, the number of species added to the red list category in this region is increasing, and the valuable genetic resources are being lost at a rapid rate. Demand for medicinal plants is increasing, and this leads to unscrupulous collection from the wild and adulteration of supplies. Providing high-quality planting material for sustainable use and thereby saving the genetic diversity of plants in the wild is important. During the last 25years of intensive research, Tropical Botanic Garden and Research Institute has developed in vitro protocol for rapid regeneration and establishment of about 40 medicinally important rare and threatened plants of Western Ghats. In situ conservation alone would not be effective in safeguarding these important species. Thus, utilizing the biotechnoligical approach to complement ex situ conservation program is becoming vital. Propagating biotechnology tools in plant conservation program is a prerequisite to succeed in sustainable use and to complement the existing ex situ measures. In addition to propagation, storage of these valuable genetic resources is equally important. In vitro slow growth of 35 species and cryopreservation using embryo/meristem/seed in 20 different species of rare medicinal plants of this region is accomplished. Plants developed in vitro of ten medicinal plants, which have restricted distribution, were reintroduced in the natural habitat as well. KeywordsConservation–Medicinal plants–Western ghats– In vitro conservation–Cryopreservation–Micropropagation
    No preview · Article · Feb 2011 · In Vitro Cellular & Developmental Biology - Plant
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    Radha R.K · Shereena S.R · Divya K · Krishnan P.N · Seeni S
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    ABSTRACT: An efficient micropropagation protocol was developed for direct shoot induction from the seedlings of Rubia cordifolia , a species sparsely distributed in the Western Ghats of India. Plant regeneration was achieved using shoot tip, nodal and split nodal half explants on Murashige and Skoog medium. Effect of plant growth regulators like 6-Benzyladenine, Kinetin, 6-Benzyladenine + Indole-3-acetic acid and 6-Benzyladenine + α-Naphthalene acetic acid on shoot multiplication and Indole-3-acetic acid, Indole-3-butyric acid and α-Naphthalene acetic acid on rooting was studied. R . cordifolia shoots showed abundant proliferation and rooting capacity, both of which are significantly influenced by the varying concentrations of the different plant growth regulators. The optimum number of shoot obtained were 5.9 and 5.2 per explants in 2 weeks on the medium supplemented with 1 mg L<SUP>-1</SUP> Benzyladenine and 0.02 mg L<SUP>-1</SUP> Indole-3-acetic acid in nodes and split vertical halves of the node respectively. The results suggest that the nodal explants (intact and split vertical halves) are better sources of shoot formation than the shoot tip explant. Shoot multiplication was rapid and consistent for 4 subcultures with 0.5 mg L<SUP>-1</SUP> Benzyladenine. The best root induction (98%) and survival was achieved on 1 mg L<SUP>-1</SUP> Indole-3-butyric acid followed by 1 mg L<SUP>-1</SUP> Indole-3-acetic acid. Rooted plantlets were successfully transferred to green house conditions with 89% survival. Micropropagated plants displayed normal phenotypes in ex situ conditions. These plantlets can be used to replenish declining populations in the wild, for the extraction of bioactive compounds and reducing pressure on wild stocks.
    Full-text · Article · Jan 2011 · International Journal of Botany
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    R K Radha · S William Decruse · P N Krishnan
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    ABSTRACT: Desiccation sensitivity and freezing tolerance of excised embryonic axes from the seeds of Nothapodytes nimmoniana (Graham) Mebberly and their cryopreservation were examined. Zygotic embryos with cotyledons excised from fully ripened fruits possessed 55.7% moisture content (MC) and 86.67% of them germinated in Murashige and Skoog (MS) medium devoid of plant growth regulators (PGR). Dehydration under laminar airflow for 120 min reduced the MC of the embryos to 19.6% and germination to 68%. Subsequent reduction in MC to 15.4, 14.2 and 12.1% also reduced the germination. Embryos dehydrated for 120 min after 1-wk-storage in Liquid nitrogen (LN) showed 60% germination, which was the optimum condition obtained in the study. The LN treated embryos developed into healthy seedlings after 30-60 d of inoculation in MS basal medium similar to desiccated control. Prolonged desiccation damaged the plumule where only root formation was observed. The study reveals the possibility of long term ex situ conservation of N. nimmoniana, which produces large and intermediate type of seeds, through zygotic embryo cryopreservation.
    Full-text · Article · Oct 2010
  • S.P. Mathew · R.K. Radha · P.N. Krishnan · S. Seeni

    No preview · Article · Apr 2010
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    ABSTRACT: Plant regeneration via somatic embryogenesis was achieved from in vitro-derived leaf base segments of Kaempferia galanga. Responses of leaf segments cultured in MS medium supplemented with Dicamba, 2,4,5-T, NAA and IAA confirmed the efficiency of synthetic auxins viz., Dicamba and 2,4,5-T to induce embryogenic callus cultures. Callus subcultured on MS medium supplemented with 0.54 or 1.07 μM NAA along with 2.22 - 6.66 μM BA produced embryoids of different stages of development on the entire surface of the callus, which on transfer to hormone-free medium developed into plantlets in 4-6 wk. Histological analysis confirmed the formation of embryoids free of vascular connection with the callus. The rooted plants were hardened and established in the field at 92% efficiency. The continued growth of the embryogenic callus through successive subculturing and the ready differentiation of the embryoids into plantlets offer reliable method for plant regeneration through somatic embryogenesis in K. galanga, which may provide an efficient and reliable system for conservation of the species.
    No preview · Article · Jul 2008 · Phytomorphology: An International Journal of Plant Morphology