Production Of Planting Material Research Articles

Above- and below-ground morpho-physiological traits indicate that biochar is a potential peat substitute for grapevine cuttings nursery production

The growing demand for grapevine planting materials, due to growing global viticulture, is promoting research studies to improve vineyard sustainability. In greenhouse nurseries, peat is the most common growing medium component used although is an expensive and non-renewable material. Indeed, the reduction of peat exploitation is receiving great attention, and currently, several materials are being investigated as peat substitutes for composing the cultivation substrates. Biochar, a carbon-rich, recalcitrant charred organic co-product of the pyrolysis or gasification process, has emerged as a potentially promising replacement for soilless substrates in nursery plant material propagation. Although several studies carried out at greenhouse nurseries have shown that biochar, can improve plant growth, only a few studies have focused on the production of grapevine plant material. To fulfil this knowledge gap and push forward the sustainability of the nursery sector, we evaluated above and below-ground morpho-physiological traits of one-year-old potted grapevine cuttings growing with 30% volume of four different biochar types (i.e., from pyrolysis and gasification) mixed with commercial peat. The present study shows that biochar can be used in growing media mixes without adverse effects on roots, improves soil water retention and leaf water potential, and improves the effects on soil microbiology.

A Temporary Immersion System as a Tool for Lowering Planting Material Production Costs Using the Example of Pennisetum × advena ‘Rubrum’

A Temporary Immersion System as a Tool for Lowering Planting Material Production Costs Using the Example of Pennisetum × advena ‘Rubrum’

Agro-technological aspects of growing branched annual apple nursery plants

The article presents an analytical review of Russian publications reporting the results of studies that search for optimal ways to affect apple plants in order to obtain branched annual plants. Since it has been proved that nursery plants of a certain age and quality are preferable for each type of intensive planting, as their use promotes the early maturity of trees thus ensuring a quick economic return on capital investments, it is currently relevant to study the production of apple tree planting material with specified biometric parameters. The use of mechanical impact on the above-ground part of grafted plants to suppress apical dominance is considered; the effectiveness of stimulating premature shoots in the crown area is confirmed. Much attention is paid by researchers to the chemical treatments of plants in the fields of fruit tree nurseries to improve the quality of annual plants. The use of cytokinin preparations and gibberellic acids is shown to be highly effective. The maximum effect on obtaining branched annual plants in most works was achieved in pomological apple varieties that are prone to the formation of premature shoots, with the budding on vegetatively propagated rootstocks of a first-grade commercial variety at a height of over 15 cm above the soil surface. One of the alternative ways of obtaining branched annual plants is wood grafting in spring and cultivation both in the field and in protected ground with a root-ball system. According to most researchers, the greatest effectiveness of the combined use of various techniques can be achieved only with strict adherence to conventional technologies for each method of cultivation.

АДАПТАЦИИ РЕГЕНЕРАНТОВ HUMULUS LUPULUS L. К УСЛОВИЯМ EX VITRO

The purpose of the study is to develop a scheme for two-stage adaptation of regenerated plants of common hop (Humulus lupulus L.) to ex vitro conditions using a hydroponic installation. Objectives: to study the influence of the method of fixing hop regenerants in a hydroponic system; to determine the duration of the adaptation stage in hydroponics; to select the substrate composition at the second stage of adaptation. The object was a sterile culture of common hop of three varieties: Tsivilsky, Flagman and Taurus, after completion of the in vitro rooting stage on a nutrient medium according to the Murashige-Skoog recipe (MS) containing glucose (20 g/l) and IBA (0.5 mg/l). The duration of the adaptation stage of hops rooted in vitro in a tidal-type hydroponic system was determined, identified by the stable maintenance of turgor by leaves and active growth of shoots – 18–21 days. The influence of the method of fixing hop regenerants in a hydroponic system was studied: the use of seedling cassettes with a substrate of perlite and vermiculite (1 : 1) showed a 25 % higher survival rate of plants compared to substrate-free cuvettes with liners made of polyethylene film. It was shown that under hydroponic installation conditions, regardless of the fixation method, the height of the shoots increased by 36.0–48.3 %. The root system of the samples adapted in cassettes with substrate was better developed and was characterized by a large number of 2nd order roots with a length of at least 4 cm. The subsequent stage of adaptation in most peat-containing substrates ensures 100 % survival and the production of high-quality planting material. Recommendations for growing hop planting material after two-stage adaptation are presented.

Effect of auxin on in vitro rooting of the triploid Musa paradisiaca L. cv. Nendran (AAB)

Banana is an important fruit crops of the family Musaceae. Tissue culture multiplication of banana is now popular and preferred due to faster multiplication, uniformly and cost-effective production of healthy planting materials. The following work was take on to study the result of various concentrations of auxin (indole-3-butyric acid-IBA). Shoots derived from sucker explants after a series of multiplication (4 to 5), regenerated plants having 4.5 cm length were dissected out and rooted in different concentrations of auxin (IBA) (0.5 to 3.5 mg/L). Root production was started after 10 days of culture and the data taken after 20 days. From all the treatment tested all media concentrations produce roots at varied level with in a period of 15-20 days of culture. Media having concentrations of IBA were used to induce in vitro rooting in plantlets of Musa cv. Nendran after six sub cultures including elongation. The elongated shoots harvested from culture bottles were transferred to MS solid media supplemented with auxin IBA at varied concentrations. The root induction rate varied from one media concentration to another. The present study revealed that IBA (1.0 mg/L) was found to be most applicable hormone for root induction in sucker shoot tip explants of Musa cv. Nendran.

Development of a Suitable in vitro Regeneration Protocol for Rahangala Pear (Pyrus pyrifolia L.) in Sri Lanka

The limitation of planting material is the major drawback to expanding pear cultivation in Sri Lanka. Hence, an in vitro protocol was developed to produce planting material for the pear variety of Rahangala. Pear shoots were used as explants and surface sterilization of explants was done using 0.5% and 1% (w/v) AgNO3 for 15 min, 5% and 10% (v/v) NaOCl for 5, 10 and 15 min. In vitro shoot multiplication and root induction were found to be suitable in full and half strengths of MS medium respectively supplemented with various concentrations of BAP (1, 1.5, 2, 2.5, 3 mg/l), IBA (0, 0.5, 1, 2, 3, 4 mg/l) and NAA (0.1 mg/l). Acclimatization was conducted in pots containing different proportions of topsoil, compost, coir dust and sand. Completely Randomized Design (CRD) was used as an experimental design for this study. During sterilization of explants, the survival rate of 86.6% was recorded after 3 weeks where the treatment was done with 1% AgNO3, and 10% NaOCl for 10 min. BAP (3.0 mg/l) produced the best shoot multiplication (1.46 ± 0.2). Significantly (p<0.05), the highest mean number of roots (1.93 ± 0.98), average length (60.6 mm), and rooting percentage (68%) were observed in ½ MS medium with 4.0 mg/l IBA. After five weeks of transplantation in pots, 100% of plantlets survived in topsoil, compost, coir dust, and sand in a ratio of 1:1:1:1 Plant Tissue Cult. & Biotech. 34(1): 25-33, 2024 (June)

Influence of growth regulators on microclonal propagation of <em>Scrophularia umbrosa</em> Dumort under <em>in vitro</em> conditions

The possibility of effective microclonal propagation of Scrophularia umbrosa Dumort for producing planting materials, aimed at preservation of the species has been shown. This will aid in the creation of artificial introduction populations of this endangered species in the Republic of Tatarstan, Russia. Optimal conditions for surface sterilization of seeds, development of microshoots, in vitro rooting and acclimatization have been developed. The most effective surface sterilization and germination was achieved with 10 % commercial bleach. Nodal explants were cultured in Murashige and Skoog’s (MS) medium with different concentrations of 6-benzylaminopurine (BAP) and indole-3-acetic acid (IAA). The maximum number of microshoots was developed on MS medium containing 1.0 mg/l of BAP and 1.0 mg/l of IAA. Full strength MS medium with only IAA or together with (indole-3-butyric acid) IBA is optimal for rapid rooting of microshoots. The combination of soil: perlite (2:1) (v/v) was the best for ex vitro acclimatization of plantlets.

The influence of the nutrient composition medium on the intensity micropropagation in vitro Fragaria × Ananassa duch.

Strawberry is one of the most economically significant crops in the world berry growing. In the course of research, the influence of the mineral composition of nutrient media on the growth and development of strawberry plants at the stage of micro-propagation proper was studied for further optimization of the technology of microclonal reproduction. The purpose of the work is to select the optimal nutrient medium for micropropagation of strawberry in vitro in the production system of healthy planting material. The research was carried out in the laboratory of biotechnology of the Russian research Institute of Fruit Grop Breeding according to generally accepted methods. The objects were important commercial varieties of garden strawberries (Fragaria × ananassa Duch.) of foreign breeding: Alba (NF 311), Darselect, Kimberly, Clery, Syria (NF 137), Florence. Variants of the nutrient media formulations used: Lee and de Fossard, Gamborg and Eveleigh (B5), Knop, Murashige-Skoog with the addition of 0.8 mg/ l cytokinin 6-BAP. Optimal recipes of the nutrient medium that promote intensive proliferation and root formation in plants have been determined. For the cultivation of the studied strawberry varieties at the stage of actual micro-propagation with a high degree of regeneration, the optimal nutrient medium is the MS medium. On the nutrient Knop’s medium, the micro-plants on the second passage acquired a reddish color, not characteristic of the studied varieties. The maximum values of plant height were noted in the control variant with MS nutrient medium in the varieties Darselect (9.1 mm) and Kimberly (8.6 mm). In all other variants of the studied nutrient media, the average height of plants did not exceed 7.7 mm. Media according to the LF and B5 recipe can be used to obtain microplants suitable for planting in ex vitro conditions, excluding the rooting stage, to accelerate the production of healthy planting material. A different reaction of strawberry varieties to the mineral composition of nutrient media has been established.

Impact of jasmonates, salicylic acid, and fungal elicitor on the health of <i>Hippeastrum hybridum</i> bulbs achieved by <i>in vitro</i> procedure

Production of high-quality vigorous plant materials obtained by micropropagation requires not only a good regeneration procedure but also efficient ways to enhance their post-transplanting ability to cope with external environmental factors. There is increasing interest in <i>Hippeastrum hybridum</i> on the horticultural market. However, the traditional method of cultivation using seeds and bulbs does not meet the growing demand and requirements of available priming procedures, thus leading to a search for more effective methods of reproduction. The aim of the study was to assess the morphogenetic abilities of bulb and flower explants for regeneration and investigate whether the enrichment of the multiplication process by exposure to elicitors will improve the health of the obtained plants through faster synthesis of phytoalexin. The study revealed that high regeneration efficiency can be obtained by using twin-scales, peduncles, and receptacles. Over 76% of explants were capable of bulblet induction and the micropropagation rates were 2.3, 2.7, and 5.0, respectively. At the rooting stage, the use of regulator-free medium was required, yielding a high percentage of rooted shoots of over 90% (shoots obtained on twin-scales and peduncles). The materials obtained <i>in vitro</i> did not differ morphologically from the initial materials. However, it was noted that, after exposure to the <i>Phoma narcissi</i> fungal elicitor or to jasmonic acid (JA) and jasmonic acid methyl ester (MeJA), the <i>in vitro</i>-derived bulbs were able to synthesize phytoalexin with higher efficiency than the untreated ones. Therefore, the described methods can be useful for <i>H. hybridum</i> multiplication. Moreover, the enrichment of the multiplication process with the stage of exposure to elicitors ensures initial immunity of <i>H. hybridum</i> to the pathogen <i>Phoma narcissi</i>, which can be used as a priming procedure.

A Comprehensive Study on the Micropropagation of Costus igneus: Media Composition, Growth, and Development

Costus igneus, commonly known as insulin plant, is an ornamental plant valued for its spiral ginger- like foliage and medicinal properties. Micropropagation is an efficient method for rapid multiplication of Costus igneus. This study aims to determine the optimal media composition and culture conditions for in vitro propagation of C. igneus. Axillary bud explants were cultured on Murashige and Skoog (MS) medium supplemented with different concentrations and combinations of plant growth regulators including auxins (indole-3-acetic acid, indole-3-butyric acid and naphthaleneacetic acid) and cytokinins (6- benzylaminopurine and kinetin). Microshoots cultured on MS + 1.5 mg/L BAP showed the highest shoot proliferation rate (98%) and maximum number of shoots per explant (12.6). Elongated microshoots were rooted on half-strength MS medium supplemented with different auxins. The highest rooting percentage (95%) and maximum number of roots per shoot (9.8) were observed on medium containing 2.5 mg/L IBA. The regenerated plantlets were acclimatized and successfully transferred to pots with 80% survival rate. Morphological and phytochemical analysis showed no significant differences between the in vitro propagated and mother plants. This study demonstrates the potential of micropropagation for large scale production of quality planting material of C. igneus. Further studies on genetic and epigenetic stability are recommended to validate this protocol for commercial applications.

In Vitro Shoot Multiplication and Rooting of 'Kashan' and 'Hervy Azerbaijan' Damask Rose (Rosa damascena Mill.) Genotypes for Cosmetic and Ornamental Applications.

The damask rose (Rosa damascena Mill.) is an ornamental-medicinal plant from the Rosaceae family, and its aromatic compounds and essential oils are applied globally in the food, cosmetic, and pharmaceutical industries. Due to its economic value, this research aimed to establish a protocol for an efficient, rapid, and cost-effective method for in vitro shoot multiplication and rooting of the R. damascena 'Kashan' and 'Hervy Azerbaijan' genotypes. Nodal segments (as primary explants) were cultured on the Murashige and Skoog (MS) medium with combinations of various plant growth regulators (PGRs) such as gibberellic acid (GA3), 6-benzylaminopurine (BAP), and indole-3-butyric acid (IBA), as well as a PGR-like substance, phloroglucinol (PG), vitamins such as ascorbic acid (AA), and activated carbon in the form of active charcoal (AC). For the establishment stage, 0.1 mg·L-1 PG, 0.2 mg·L-1 GA3, and 1 mg·L-1 BAP were added to the media. Secondary explants (nodal segments containing axillary buds produced from primary explants) were obtained after 30 days of in vitro culture and transferred to the proliferation media supplemented with different concentrations of BAP (0, 0.5, 1, 1.5, 2, and 2.5 mg·L-1) and GA3 (0, 0.1, 0.2, 0.4, 0.8, and 1 mg·L-1) together with 0.1 mg·L-1 PG and 20 mg·L-1 of AA. The rooting media were augmented with different concentrations of BAP and GA3 with 0.1 mg·L-1 of IBA, PG and 20 mg·L-1 of AA and AC. The results showed that the highest regeneration coefficient (4.29 and 4.28) and the largest number of leaves (23.33-24.33) were obtained in the explants grown on the medium supplemented with 2 mg·L-1 BAP and 0.4 mg·L-1 GA3 for the 'Kashan' and 'Hervy Azerbaijan' genotypes, respectively. Likewise, this PGR combination provided the shortest time until bud break (approximately 6.5 days) and root emergence (approximately 10 days) in both genotypes. The highest number of shoots (4.78 per explant) and roots (3.96) was achieved in this medium in the 'Kashan' rose. Stem and root lengths, as well as stem and root fresh and dry weights, were also analyzed. In most measured traits, the lowest values were found in the PGRs-free control medium. Rooted plantlets were transferred to pots filled with perlite and peat moss in a 2:1 proportion and were acclimatized to ambient greenhouse conditions with a mean 90.12% survival rate. This research contributes significantly to our understanding of Damask rose propagation and has practical implications for the cosmetic and ornamental plant industries. By offering insights into the manipulation of regeneration processes, our study opens up new possibilities for the effective production of high-quality plant material.

In vitro microrhizome induction in Acorus calamus L., a commercially important aromatic medicinal plant

Acorus calamus L., is a leading aromatic medicinal plant that produces pungent aromatic rhizomes that are valued worldwide as an important herbal medicine and is one of the main ingredients in several polyherbal formulations used for neurological and metabolic disorders. The present investigation aims to develop an efficient protocol for in vitro microrhizome induction in A. calamus for the large-scale production of disease-free planting material for commercial purposes. In vitro derived shoots were initiated on MS medium supplemented with different concentrations of sucrose alone (3–10 %) and different concentrations of sucrose (6 % and 7 %) with varying concentrations of BAP alone (0.5–2 mg/L) and combinations with IAA (0.5 and 1 mg/L) and NAA (0.5 and 1 mg/L) were carried out for the microrhizome induction experiments. In different concentrations of sucrose used, healthy and disease-free microshoots of A. calamus were obtained in MS media supplemented with 7 % sucrose followed by 6 % sucrose. The highest shoot length (24.79 ± 0.03 cm) and microrhizome size (4.98 ± 0.03 cm length and 399.60 ± 0.37 mg fresh weight) were obtained in MS solid medium supplemented with 1 mg/L BAP and 0.5 mg/L NAA with 7 % sucrose followed by the same hormone concentration with 6 % sucrose (shoot length-23.68 ± 0.03 cm, microrhizome length- 4.68 ± 0.03 cm and fresh weight of microrhizome-376.60 ± 0.57 mg). The developed protocol can be used for large-scale production of disease-free propagules without rooting and acclimatization and enhance the production of true-to-type planting material.

Transcriptional and physio-chemical responses of Tectona grandis L. triggered by teak defoliator

Tectona grandis (L) is a popular timber species cultivated in the Indian sub-continent and other countries because of its economic significance. However, the lack of quality planting material with special reference to defoliator herbivory is a major constraint because it negatively influences the establishment of thriving plantations. To address this, the so-called pest response in teak seedlings was investigated at the biochemical and molecular levels. The stress was levied by allowing defoliator infestation on 1-year-old seedlings for nine days at 3-day intervals, whereas control plants were maintained as uninfected. A considerable difference was observed for the considered traits among the infested plants at different intervals compared with the control. Total soluble protein, soluble sugars, tannin, and lignin accumulation increased with increasing stress intensity. Furthermore, catalase, polyphenol oxidase, and phenol content increased after defoliator feeding, whereas peroxidase activity decreased compared with control plants. In addition, six herbivory stress-responsive genes revealed the molecular response of teak. Among them, four genes viz. zinc finger protein (ZFP2), ethylene-responsive factor (ERF-1), catalase-1 (Cat1) and peroxidase super family protein (Psfp) were highly expressed in the infected plants as compared with the control plants. In contrast, the MYB-DNA binding domain gene was downregulated in infected plants compared to uninfected plants. Cytochrome P450 was up-regulated in T1 and T2 plants, while it was down-regulated in T3. Genes were highly expressed as the stress frequency increased. This study offers an improved understanding of biochemical and transcriptional defense responses against the teak defoliator, which could further support teak improvement for quality planting material production.

Participatory research processes for the production of plant material in food security context

The production of quality seed constitutes one of the main challenges for food security and sovereignty of both urban and rural populations, requiring processes that recognize not only the potential of the territories but also the diversity associated withpeasant communities and their forms of production. , so as to reduce the gaps resulting from the use of seeds of low or no physical, genetic, physiological and sanitary quality that reduce the impact on the yields of banana, pineapple and cassava crops. To this end, a participatory research process was carried out to meet the demand for projects that improve the food and nutritional security of the population of the department of Casanare -Colombia. As a result, the methodology was implemented based on meetings with producers of productive systems for the construction of knowledge from social cartography, the preparation of economic calendars and mappings of actors and needs. This reading of the environments allowed the development of activities for the participatory selection of planting materials, processes of intensive multiplication of materials, as well as the transformation and production of banana, pineapple and cassava derivatives. The conclusions have to do with the importance of considering the producer as an active participant in the research processes in the construction of knowledge that contributes to the generation and appropriation of solutions based on science, technology and innovation in favor of food security and with this the reduction of poverty. In the near future, it is planned to carry out a training process for trainers with allied universities to leave installed capacities and to promote access and availability of quality seed.

Silver nanoparticle-induced in vitro flowering in Dendrocalamus strictus (Roxb.) nees and genetic fidelity assessment of regenerants using molecular markers.

Dendrocalamus strictus (Roxb.) Nees, generally referred to as 'Male bamboo,' is a globally prevalent and highly significant species of bamboo. It is a versatile species and possesses notable industrial significance. However, despite its numerous applications, the production of this plant is insufficient to fulfill the worldwide demand. The challenges that impede the dissemination of D. strictus encompass the unpredictable blooming pattern (30-70 years), low seed production, and limited seed viability. Therefore, tissue culture presents a reliable and effective option for the mass production of standardized planting material. This study investigated the effects of silver nanoparticles (AgNPs) at a concentration of 6.0 mg L- 1 in the Murashige and Skoog (MS) nutrient medium fortified with pre-optimized plant growth regulators (3.0 mg L- 1 6-benzylaminopurine + 0.5 mg L- 1 α-naphthalene acetic acid) on the induction of flowering in a controlled environment in D. strictus. The use of AgNPs in the media induced a maximum of 14 inflorescences per culture vessel, 9 flowers per inflorescence, and improved the performance of the micropropagated plantlets during acclimatization in the greenhouse and field. The ISSR and SCoT amplified polymorphic DNA analysis of the regenerants resulted in the formation of 49 bands (300 to 2000bp size) and 36 scorable bands (350 to 2000bp) respectively. All the PCR amplicons produced by SCoT and ISSR were monomorphic confirming the genetic uniformity of the tissue cultured plants of D. strictus with the mother plant. It can be inferred that the incorporation of AgNPs during the shoot proliferation phase has the potential to stimulate in vitro flowering in D. strictus. This finding could provide valuable insights into innovative strategies for enhancing crop productivity and genetic manipulation for accelerated breeding and agricultural advancement.

Production of quality planting material in commercial banana cvs. Rasthali (Silk, AAB) and Neypoovan (Neypoovan, AB) through farmer-friendly macropropagation technique and their field evaluation

Macropropagation is a low cost farmer friendly alternative to tissue culture propagation to cater to the farmers’ need for quality planting material at an affordable cost. Attempts were made to study the effect of age and hot water treatment on macropropagation of cvs. Rasthali (AAB, Silk) and Neypoovan (AB, Neypoovan) in pot and bed methods. Age of the sucker, hot water treatment and propagation method played a vital role on shoot bud production in both the test cultivars. In cv. Rasthali, bed method produced maximum number of plants (50.5) in five months old suckers subjected to hot water treatment in 42.2 days but the same treatment produced only 24.7 plants under pot method. In cv. Neypoovan, five months old suckers subjected to hot water treatment resulted in 23.3 and 16.7 plants in bed and pot methods respectively. Therefore, five months old suckers with hot water treatment under bed method are more appropriate for macropropagation purpose in banana. Results of field evaluation indicated that macropropagated bananas were at par with control in respect of all plant parameters like plant height, bunch weight and fruit parameters like total soluble sugar (TSS) and acidity.

From Lab to Bouquet: The Biotechnological Frontier in Modern Floriculture for Sustainable and Resilient Flower Farming

Modern floriculture faces unprecedented challenges in meeting the rising global demand for flowers while addressing environmental concerns and the need for sustainability. This paper explores the transformative role of biotechnology in revolutionizing flower farming practices, offering solutions for sustainable and resilient cultivation. The integration of biotechnological tools such as genetic engineering, tissue culture, and molecular breeding techniques has opened new avenues for enhancing the efficiency and resilience of flower crops. Genetic modification allows the development of varieties with improved traits, such as disease resistance, longer vase life, and enhanced stress tolerance. Tissue culture facilitates the rapid propagation of elite plant material, ensuring consistent and high-quality flower production. Furthermore, molecular breeding enables the precise selection of desirable traits, accelerating the traditional breeding process. The use of molecular markers for disease resistance, flower colour, and fragrance can streamline the development of new flower varieties tailored to market preferences. Additionally, advanced omics technologies, including genomics, transcriptomic, and metabolomics, provide valuable insights into the molecular mechanisms underlying flower development and responses to environmental stress. Biotechnological interventions extend beyond genetic improvements, encompassing sustainable cultivation practices. Precision agriculture, enabled by sensor technologies and data analytics, optimizes resource utilization, minimizing environmental impact. Smart irrigation systems, nutrient management, and pest control strategies contribute to the efficient use of resources and reduced ecological footprint. The biotechnological frontier in modern floriculture not only addresses current challenges but also positions the industry to adapt to future uncertainties. Climate change, emerging pests, and shifting market demands necessitate resilient flower farming systems. Biotechnology offers the tools to develop adaptable flower varieties and sustainable production methods, promoting long-term ecological and economic viability.

Effect of Growing Media and Cow Urine on Seedling Growth, Leaf Nutrient Status and Economics of Papaya (Carica papaya L.)

In recent decades, research attention is focusing on utilize animal wastes for the quality planting material production with the various growing media. In this study, a pot experiment was conducted on papaya cv. Arka Surya at the nursery unit of the Department of Horticulture, Agricultural University of Kota, Rajasthan to assessment the impact of growing media and organic liquid (cow urine) on seedling growth, leaf nutrient status and economic attributes. This experiment was laid out in a factorial complete randomized design with two factors consisting eleven growing media and two levels of cow urine (with or without). Among different treatments combination used the medium Soil + Sand + Vermiculite + Cocopeat + Perlite (1:1:1:1:1) with cow urine found significantly superior over other treatments and control with respect to maximum leaf area (51.85 cm2), vigour index-I (1550.27 cm), vigour index-II (59.55 g), fresh weight of seedling (2.86 g) and dry weight of seedling (0.42 g) at 75 days after sowing. The highest number of secondary roots (17.00) were also found in the same treatment as compared to rest of the treatments. Further, it was also found superior with relation to maximum nitrogen (1.87%), phosphorus (1.14%) and potassium (2.26%) content. The highest number of stout and healthy seedlings (9000) per 250 g seeds. Therefore, maximum total gross return (₹ 90000) obtained by selling of these seedlings with higher net income (₹ 17363) due to treatment T10-Soil + Sand + Vermiculite + Cocopeat + Perlite (1:1:1:1:1) with cow urine over control. However, it is recommended that Soil + Sand + Vermiculite + Cocopeat + Perlite (1:1:1:1:1) with cow urine should be used for healthier growth and higher nutrient quality with more net profitability.

Development of Simplified Recombinase Polymerase Amplification Assay for Rapid and Robust Detection of Citrus Yellow Vein Clearing Virus.

Citrus is an economically important fruit crop, belongs to family Rutaceae, cultivated commercially in over 130 countries, which holds a leading profitable position in the international market. The most important citrus varieties are mandarins, oranges, lemons, sweet limes, grapefruits and pomelos. Citrus yellow vein clearing virus (CYVCV) is an important graft transmissible plant pathogen known to reduce productivity of citrus fruits due to its predominant association and widespread occurrence. Requirement of fast, reliable, efficient & economical CYVCV indexing assay is a prerequisite for production of healthy planting material. Currently, nucleic acid isolation and thermal cycler-based assay available for CYVCV indexing is a cumbersome lab intensive method. The present study was undertaken to develop and validate reverse transcription-recombinase polymerase amplification (RT-RPA) assay requiring no tedious RNA isolation, separate cDNA synthesis and costlier instrument like thermo-cycler. Optimized RT-RPA assay was able to amplify CYVCV up to 10-7 dilution (equivalent to 0.1pg/μl) with the prepared templates of both RNA and crude saps and showed higher sensitivity in detection of CYVCV infection in field samples as compared to the conventional RT-PCR. Developed RT-RPA assay showed high specificity without any cross-reaction with other citrus pathogens (Indian citrus ringspot virus, citrus yellow mosaic virus, citrus tristeza virus, citrus exocortis viroid and huanglongbing). RT-RPA using crude leaf sap as template is quite simple, robust, highly sensitive, time and cost effective; therefore, it can be used in resource constrained laboratories as screening tool, for field surveys and on-site testing programs in farms, nurseries and biosecurity. Present study, first time reports the development, optimization and validation of crude sap-based RT-RPA assay for the detection of CYVCV infection in citrus plants namely; Kinnow mandarin, Mosambi and Grape fruit.

Development of Polyclonal Antibodies-Based Serological Method for the Detection of Calanthe Mild Mosaic Virus and Application in Virus Certification Programme.

Calanthe mild mosaic virus (CalMMV) infecting orchids is an important potyvirus which is known to cause mild leaf mosaic and flower colour-breaking symptoms in Calanthe and other orchid plants. The present study reports the production of polyclonal antibodies against CalMMV using bacterially expressed recombinant coat protein as immunogen, which in turn would be useful in routine indexing and screening of orchid germplasm. The coat protein (CP) gene (~ 807bp) of CalMMV isolated from infected orchid sample was cloned in expression vector, pET-28a ( +) that yielded ~ 31kDa fusion protein with Histidine tag (His6BP). The expression of fusion CP was confirmed through SDS-PAGE and Western blotting. The His6BP-CalMMV-CP obtained in soluble state after purification was used to immunize New Zealand white rabbit for the production of polyclonal antibodies (PAb). The PAb produced against the purified fusion protein successfully detected CAlMMV in the orchid samples at a dilution of 1:2000 in direct antigen-coated enzyme-linked immunosorbent assay (DAC-ELISA). This study presents the first report of Histidine tag (His6BP) fusion CalMMV-CP-based antibody production and its successful application in the identification of the virus in orchid plants. Outcome of this study will be helpful in routine certification programmes, screening of orchid germplasm and production of CalMMV-free planting materials of orchids.