Agrochemical-free genetically modified and genome-edited crops: Towards achieving the United Nations sustainable development goals and a 'greener' green revolution.

Sustainable farming on ever-shrinking agricultural land and declining water resources for the growing human population is one of the greatest environmental and food security challenges of the 21st century. Conventional, age-old organic farming practices alone, and foods based on costly cellular agriculture, do not have the potential to be upscaled to meet the food supply challenges for feeding large populations. Additionally, agricultural practices relying on chemical inputs have a well-documented detrimental impact on human health and the environment. As the available farming methods have reached their productivity limits, new approaches to agriculture, combining friendly, age-old farming practices with modern technologies that exclude chemical interventions, are necessary to address the food production challenges. Growing genetically modified (GM) crops without chemical inputs can allow agricultural intensification with reduced adverse health and environmental impacts. Additionally, integrating high-value pleiotropic genes in their genetic improvement coupled with the use of modern agricultural technologies, like robotics and artificial intelligence (AI), will further improve productivity. Such 'organic-GM' crops will offer consumers healthy, agrochemical-free GM produce. We believe these agricultural practices will lead to the beginning of a potentially new chemical-free GM agricultural revolution in the era of Agriculture 4.0 and help meet the targets of the United Nations Sustainable Development Goals (SDGs). Furthermore, given the advancement in the genome editing (GE) toolbox, we ought to develop a new category of 'trait-reversible GM crops' to avert the fears of those who believe in ecological damage by GM crops. Thus, in this article, we advocate farming with no or minimal chemical use by combining chemical-free organic farming with the existing biofortified and multiple stress tolerant GM crops, while focusing on the development of novel 'biofertilizer-responsive GE crops' and 'trait-reversible GE crops' for the future.

Ameliorating the effects of multiple stresses on agronomic traits in crops: modern biotechnological and omics approaches.

While global climate change poses a significant environmental threat to agriculture, the increasing population is another big challenge to food security. To address this, developing crop varieties with increased productivity and tolerance to biotic and abiotic stresses is crucial. Breeders must identify traits to ensure higher and consistent yields under inconsistent environmental challenges, possess resilience against emerging biotic and abiotic stresses and satisfy customer demands for safer and more nutritious meals. With the advent of omics-based technologies, molecular tools are now integrated with breeding to understand the molecular genetics of genotype-based traits and develop better climate-smart crops. The rapid development of omics technologies offers an opportunity to generate novel datasets for crop species. Identifying genes and pathways responsible for significant agronomic traits has been made possible by integrating omics data with genetic and phenotypic information. This paper discusses the importance and use of omics-based strategies, including genomics, transcriptomics, proteomics and phenomics, for agricultural and horticultural crop improvement, which aligns with developing better adaptability in these crop species to the changing climate conditions.

Quality trait improvement in horticultural crops: OMICS and modern biotechnological approaches.

Horticultural crops are an essential part of food and nutritional security. Moreover, these form an integral part of the agricultural economy and have enormous economic potential. They are a rich source of nutrients that are beneficial to human health. Plant breeding of horticultural crops has focussed primarily on increasing the productivity and related traits of these crops. However, fruit and vegetable quality is paramount to their perishability, marketability, and consumer acceptance. The improved nutritional value is beneficial to underprivileged and undernourished communities. Due to a declining genetic base, conventional plant breeding does not contribute much to quality improvement as the existing natural allelic variations and crossing barriers between cultivated and wild species limit it. Over the past two decades, 'omics' and modern biotechnological approaches have made it possible to decode the complex genomes of crop plants, assign functions to the otherwise many unknown genes, and develop genome-wide DNA markers. Genetic engineering has enabled the validation of these genes and the introduction of crucial agronomic traits influencing various quality parameters directly or indirectly. This review discusses the significant advances in the quality improvement of horticultural crops, including shelf life, aroma, browning, nutritional value, colour, and many other related traits.

Trait specific marker-based characterization and population structure analysis in rice (Oryza sativa L.) germplasm of Kashmir Himalayas.

BACKGROUND: Rice is a key food grain contributor to the global food grain basket and is considered the main food crop in India with a large number of varieties released every year. SSR markers have proven to be an excellent tool for studying genetic diversity. As a result, the present study was done to characterize and assess genetic diversity as well as population structural aspects. METHODS AND RESULTS: Fifty genotypes of rice were characterized using 40 SSR markers to assess the genetic diversity and genetic relationship. A total of 114 alleles were amplified with an average of 2.85 alleles per locus. The Polymorphism Information Content (PIC) values varied from 0.30 (RM162) to 0.58 (RM413) with an average of 0.44. Gene diversity was in the range of 0.35 (RM162) to 0.66 (RM413), with an average value of 0.52, while heterozygosity ranged from 0.18 (RM27) to 0.74 (RM55), with an average of 0.39. The population structure revealed a narrow genetic base with only three major subpopulations. Analysis of molecular variance revealed that 74% of the variation was attributed within individuals, 23% was among individuals, and 3% was among populations. Pairwise Fst value of population A & B is 0.024, population B & C is 0.120 and population A & C is 0.115. Dendrogram grouped the genotypes into three clusters with wide variation among the accessions. CONCLUSION: Genotyping combined with phylogeny and population structure analysis proved to be a powerful method for characterizing germplasm in this study. There is significant gene flow within populations, as well as the presence of different combinations of alleles, and that allelic exchange rates within the populations are higher than among the populations. Assessing the genetic diversity among individual genotypes within populations is quite useful in selecting candidate parents for future breeding programs to improve the target traits in rice for the Himalayan region.

Robotics-assisted, organic agricultural-biotechnology based environment-friendly healthy food option: Beyond the binary of GM versus Organic crops.

Human society cannot afford the luxury of the business-as-usual approach when dealing with the emerging challenges of the 21st century. The challenges of food production to meet the pace of population growth in an environmentally-sustainable manner have increased considerably, emphasizing the need to explore newer approaches to agriculture. Agrochemical-based agricultural practices are known to have serious environmental and health implications. Even conventional organic farming is not sustainable in the long run. Although some "age-old" practices are useful, these will not help feed more people on the same or less land more sustainably. Sustainable intensification is the way forward. There is a need to incorporate a customer-centric outlook and make the organic system sustainable. Here, we bring forth the necessity to enhance the efficiency of organic agriculture by the inclusion of robotics and agrochemical-free GM seeds. Such an organic-GM hybrid agriculture system integrated with the use of artificial intelligence (AI) based technologies will have better energy efficiency. The produce from such a system will offer consumers a 'third' choice and create a new food label, 'organically-grown GM produce'.

Callus induction and regeneration in high-altitude Himalayan rice genotype SR4 via seed explant.

SR4 genotype of rice is high altitude Himalayan rice prone to various abiotic stresses such as cold stress and therefore gives a poor yield. An efficient protocol for callusing and regeneration via direct and indirect means was established using mature seeds as an explant which can be utilized for molecular studies for genetic advancement of Himalayan rice genotype SR4 through transformation. Highest frequency (96.6%) of callus induction was obtained on MS media 3.0 mg/L 2, 4-D. While maximum regeneration frequency (100%), number of shoots with maximum length 9.14 ± 0.204 (cm) from callus was recovered from MS media amended with 5.0 mg/L BAP in combination with 0.5 mg/L NAA with highest number of shoots having an average shoot length 9.14 ± 0.204 (cm) after four weeks of culture. Direct multiple shoot regeneration from seed explants was obtained using various concentrations of TDZ and BAP with highest regeneration frequency was observed on MS media fortified with 6 mg/L of TDZ with maximum number of shoots. The shoots developed roots on MS media supplemented with 0.6 mg/L IBA.

The menace of saffron adulteration: Low-cost rapid identification of fake look-alike saffron using Foldscope and machine learning technology.

Saffron authenticity is important for the saffron industry, consumers, food industry, and regulatory agencies. Herein we describe a combo of two novel methods to distinguish genuine saffron from fake in a user-friendly manner and without sophisticated instruments. A smartphone coupled with Foldscope was used to visualize characteristic features and distinguish "genuine" saffron from "fake." Furthermore, destaining and staining agents were used to study the staining patterns. Toluidine blue staining pattern was distinct and easier to use as it stained the papillae and the margins deep purple, while its stain is lighter yellowish green toward the central axis. Further to automate the process, we tested and compared different machine learning-based classification approaches for performing the automated saffron classification into genuine or fake. We demonstrated that the deep learning-based models are efficient in learning the morphological features and classifying samples as either fake or genuine, making it much easier for end-users. This approach performed much better than conventional machine learning approaches (random forest and SVM), and the model achieved an accuracy of 99.5% and a precision of 99.3% on the test dataset. The process has increased the robustness and reliability of authenticating saffron samples. This is the first study that describes a customer-centric frugal science-based approach to creating an automated app to detect adulteration. Furthermore, a survey was conducted to assess saffron adulteration and quality. It revealed that only 40% of samples belonged to ISO Category I, while the average adulteration percentage in the remaining samples was 36.25%. After discarding the adulterants from crude samples, their quality parameters improved significantly, elevating these from ISO category III to Category II. Conversely, it also means that Categories II and III saffron are more prone to and favored for adulteration by fraudsters.

Polyphenolics, antioxidant characterization and DNA barcoding of Kala zeera [Bunium persicum (Boiss.) Fedtsch] through multiple barcode analysis to unravel best barcode combination.

BACKGROUND: Kala zeera [Bunium persicum (Boiss.) Fedtsch] is one of the important spice crops of North Western Himalayas with lot of medicinal and culinary values. In spite of having great importance, this crop is under the threat of extinction due to loss of habitat and lack of awareness. The limited availability of the seeds has ultimately increased the economic value of this spice. The upmarket of Kala zeera leads to its adulteration with other black seeds and cumin seeds. The present investigation was undertaken to evaluate polyphenolics and antioxidant properties of Kala zeera genotypes collected from North Western Himalayas and to develop DNA barcodes that can ensure their purity and can also guide in conservation of selected Kala zeera germplasm lines. METHODS AND RESULTS: Various locations of North Western Himalayas were explored for collecting 31 diverse germplasm lines of Kala zeera. The collected germplasm was maintained at our experimental stations during 2019-2020 and 2020-2021. These genotypes were evaluated for different seed traits and the methanolic extract from Kala zeera seeds was examined for total phenolic content, total flavonoid content, antioxidant activities by DPPH and FRAP. The results revealed significant variation in seed traits, polyphenolic content and antioxidant properties. 100 seed weight ranged from 0.05 to 0.35 g, TPC ranged from 7.5 to 22.56 mg/g, TFC ranged from 0.58 to 4.15 mg/g, antioxidant properties DPPH ranged from 168 to 624.4 µg/ml and FRAP ranged from 0.72 to 6.91 mg/g. Further, three different barcodes (ITS, rbcL and psbA-trnH) were used to reveal the authenticity of selected Kala zeera. MEGA 5 software was used for clustering and the barcodes did clustering based on geographical distribution of Kala zeera germplasm. CONCLUSION: Based on molecular barcoding, best barcode combination was identified that may discriminate the Kala zeera germplasm vis-a-vis can authenticate their purity. Moreover, the identified DNA barcodes will have significant role in studying the evolutionary biology of Bunium species and will be important for designing a strategy to conserve the selected Kala zeera germplasm lines. The identified genotypes with high phenolic content and antioxidant activity can further be utilized in Kala zeera breeding programmes.

Understanding saffron biology using omics- and bioinformatics tools: stepping towards a better Crocus phenome.

Saffron is a unique plant in many aspects, and its cellular processes are regulated at multiple levels. The genetic makeup in the form of eight chromosome triplets (2n = 3x = 24) with a haploid genetic content (genome size) of 3.45 Gbp is decoded into different types of RNA by transcription. The RNA then translates into peptides and functional proteins, sometimes involving post-translational modifications too. The interactions of the genome, transcriptome, proteome and other regulatory molecules ultimately result in the complex set of primary and secondary metabolites of saffron metabolome. These complex interactions manifest in the form of a set of traits 'phenome' peculiar to saffron. The phenome responds to the environmental changes occurring in and around saffron and modify its response in respect of growth, development, disease response, stigma quality, apocarotenoid biosynthesis, and other processes. Understanding these complex relations between different yet interconnected biological activities is quite challenging in saffron where classical genetics has a very limited role owing to its sterility, and the absence of a whole-genome sequence. Omics-based technologies are immensely helpful in overcoming these limitations and developing a better understanding of saffron biology. In addition to creating a comprehensive picture of the molecular mechanisms involved in apocarotenoid synthesis, stigma biogenesis, corm activity, and flower development, omics-technologies will ultimately lead to the engineering of saffron plants with improved phenome.

High-value pleiotropic genes for developing multiple stress-tolerant biofortified crops for 21st-century challenges.

The agriculture-based livelihood systems that are already vulnerable due to multiple challenges face immediate risk of increased crop failures due to weather vagaries. As breeders and biotechnologists, our strategy is to advance and innovate breeding for weather-proofing crops. Plant stress tolerance is a genetically complex trait. Additionally, crops rarely face a single type of stress in isolation, and it is difficult for plants to deal with multiple stresses simultaneously. One of the most helpful approaches to creating stress-resilient crops is genome editing and trans- or cis-genesis. Out of hundreds of stress-responsive genes, many have been used to impart tolerance against a particular stress factor, while a few used in combination for gene pyramiding against multiple stresses. However, a better approach would be to use multi-role pleiotropic genes that enable plants to adapt to numerous environmental stresses simultaneously. Herein we attempt to integrate and present the scattered information published in the past three decades about these pleiotropic genes for crop improvement and remodeling future cropping systems. Research articles validating functional roles of genes in transgenic plants were used to create groups of multi-role pleiotropic genes that could be candidate genes for developing weather-proof crop varieties. These biotech crop varieties will help create 'high-value farms' to meet the goal of a sustainable increase in global food productivity and stabilize food prices by ensuring a fluctuation-free assured food supply. It could also help create a gene repository through artificial gene synthesis for 'resilient high-value food production' for the 21st century.

Saffron: A potential drug-supplement for severe acute respiratory syndrome coronavirus (COVID) management.

Severe acute respiratory syndrome coronavirus 2, SARS-CoV-2 (COVID-19), came as a significant health care challenge for humans in 2019-20. Based on recent laboratory and epidemiological studies, a growing list of mutations in the virus has the potential to enhance its transmission or help it evade the immune response. To further compound the problems, there are considerable challenges to the availability of effective, affordable, safe vaccines on a mass scale. These impediments have led some to explore additional options available in traditional medicines, especially immune-boosting natural products. Saffron has been used for centuries to treat fever, bronchitis, cold and other immune, respiratory disorders. Herein, we discuss the potential role of saffron during and after COVID-19 infection, focusing on immunomodulation, respiratory, renal, and cardiovascular functions. As a nutraceutical or drug supplement, it can alleviate the magnitude of COVID-19 symptoms in patients. The anti-inflammatory, antioxidant, and other medicinal properties attributed to saffron bioactive compounds can help in both pre-and post-infection management strategies. The abnormalities associated with COVID-19 survivors include anxiety, depression, sleep disturbances, and post-traumatic stress disorder. Saffron can help manage these post-hospitalization abnormalities (sub-acute and chronic) too, owing to its anti-depressant property. It can help common people boost immunity and manage depression, stress and anxiety caused due to prolonged lockdown, isolation or quarantine.

Differential Bioaccumulation of Select Heavy Metals from Wastewater by Lemna minor.

The capacity of Lemna minor to remediate toxic heavy metals from wastewater is reasonably well documented. In view of the pivotal role of this species in the environmental clean-up, here we evaluated the bioaccumulation potential of L. minor for cadmium (Cd), lead (Pb), and nickel (Ni) through a controlled experiment. L. minor tolerated the metals Cd, Ni, and Pb up to 0.5, 5, and 8 mg/L, respectively, and beyond these concentrations the toxicity symptoms appeared. Bio-concentration factor varied at different concentrations of heavy metals tested. Overall, L. minor showed good phytoremediation potential for all the three tested heavy metals (Cd, Ni, and Pb), though in relative terms it was more effective in extracting Ni and Cd, as compared to Pb, both in single and mixed concentrations. In view of the growing pollution in Kashmir Himalayan aquatic habitats the phytoremediation by invasive species such as L. minor promises to be one of the best choices than other native plants for cleaning up of polluted soils/water because of its fast growth rate, high abundance, easy handling, and wide distribution in Kashmir Himalayan aquatic ecosystems.

Time to Redefine Organic Agriculture: Can't GM Crops Be Certified as Organics?

The challenges of sustainable food production without damaging the environment for a growing human population have increased considerably. The current agricultural practices involving chemical fertilizers and even organic farming are not sustainable in the long run and can have deleterious effects on the environment. Thus, new, innovative solutions need to be identified and propagated for tackling this. Among such innovations, that can complement conventional as well as organic farming methods, are genetic modification (GM) and aquaculture. Yet, GM technologies often face resistance from civil groups owing to an 'unknown' fear, akin to Frankenstein's monster. How real is this fear? Our discussion rests on basic questions like, why can't 'organics' include GM crops that do not require chemical inputs for cultivation, and can GM crops like Golden rice qualify to be 'organic' if cultivated through organic practices? Do we need to rethink organic agriculture in the context of the present and future challenges of 21st century?

Multiplex Fluorescent, Activity-Based Protein Profiling Identifies Active α-Glycosidases and Other Hydrolases in Plants.

With nearly 140 α-glycosidases in 14 different families, plants are well equipped with enzymes that can break the α-glucosidic bonds in a large diversity of molecules. Here, we introduce activity-based protein profiling (ABPP) of α-glycosidases in plants using α-configured cyclophellitol aziridine probes carrying various fluorophores or biotin. In Arabidopsis (Arabidopsis thaliana), these probes label members of the GH31 family of glycosyl hydrolases, including endoplasmic reticulum-resident α-glucosidase-II Radial Swelling3/Priority for Sweet Life5 (RSW3/PSL5) and Golgi-resident α-mannosidase-II Hybrid Glycosylation1 (HGL1), both of which trim N-glycans on glycoproteins. We detected the active state of extracellular α-glycosidases such as α-xylosidase XYL1, which acts on xyloglucans in the cell wall to promote cell expansion, and α-glucosidase AGLU1, which acts in starch hydrolysis and can suppress fungal invasion. Labeling of α-glycosidases generates pH-dependent signals that can be suppressed by α-glycosidase inhibitors in a broad range of plant species. To demonstrate its use on a nonmodel plant species, we applied ABPP on saffron crocus (Crocus sativus), a cash crop for the production of saffron spice. Using a combination of biotinylated glycosidase probes, we identified and quantified 67 active glycosidases in saffron crocus stigma, of which 10 are differentially active. We also uncovered massive changes in hydrolase activities in the corms upon infection with Fusarium oxysporum using multiplex fluorescence labeling in combination with probes for serine hydrolases and cysteine proteases. These experiments demonstrate the ease with which active α-glycosidases and other hydrolases can be analyzed through ABPP in model and nonmodel plants.

Host-Pathogen Interaction in Fusarium oxysporum Infections: Where Do We Stand?

Fusarium oxysporum, a ubiquitous soilborne pathogen, causes devastating vascular wilt in more than 100 plant species and ranks 5th among the top 10 fungal plant pathogens. It has emerged as a human pathogen, too, causing infections in immune-compromised patients. Therefore, it is important to gain insight into the molecular processes involved in the pathogenesis of this transkingdom pathogen. A complex network comprising interconnected and overlapping signal pathways-mitogen-activated protein kinase signaling pathways, Ras proteins, G-protein signaling components and their downstream pathways, components of the velvet (LaeA/VeA/VelB) complex, and cAMP pathways-is involved in perceiving the host. This network regulates the expression of various pathogenicity genes. However, plants have evolved an elaborate protection system to combat this attack. They, too, possess intricate mechanisms at the molecular level which, once triggered by pathogen attack, transduce signals to activate defense response. This review focuses on understanding and presenting a wholistic picture of the molecular mechanisms of F. oxysporum-host interactions in plant immunity.

SSR based genetic diversity of pigmented and aromatic rice (Oryza sativa L.) genotypes of the western Himalayan region of India.

A set of 24 of SSR markers were used to estimate the genetic diversity in 16 rice genotypes found in Western Himalayas of Kashmir and Himachal Pradesh, India. The level of polymorphism among the genotypes of rice was evaluated from the number of alleles and PIC value for each of the 24 SSR loci. A total of 68 alleles were detected across the 16 genotypes through the use of these 24 SSR markers The number of alleles per locus generated varied from 2 (RM 338, RM 452, RM 171) to 6 (RM 585, RM 249, RM 481, RM 162). The PIC values varied from 0.36 (RM 1) to 0.86 (RM 249) with an average of 0.62 per locus. Based on information generated, the genotypes got separated in six different clusters. Cluster 1 comprised of 4 genotypes viz; Zag 1, Zag 13, Pusa sugandh 3, and Zag 14, separated from each other at a similarity value of 0.40. Cluster second comprised of 3 landraces viz; Zag 2. Zag 4 and Zag10 separated from each other at a similarity value of 0.45. Cluster third comprised of 3 genotypes viz; Grey rice, Mushk budji and Kamad separated from each other at a similarity value of 0.46. Cluster fourth had 2 landraces viz; Kawa kreed and Loual anzul, and was not sub clustered. Fifth cluster had 3 genotypes viz; Zag 12, Purple rice and Jhelum separated from each other at a similarity value of 0.28. Cluster 6 comprised of a single popular variety i.e. Shalimar rice 1 with independent lineage.

Challenges of climate change: omics-based biology of saffron plants and organic agricultural biotechnology for sustainable saffron production.

Kashmir Valley is a major saffron (Crocus sativus Kashmirianus) growing area of the world, second only to Iran in terms of production. In Kashmir, saffron is grown on uplands (termed in the local language as "Karewas"), which are lacustrine deposits located at an altitude of 1585 to 1677 m above mean sea level (amsl), under temperate climatic conditions. Kashmir, despite being one of the oldest historical saffron-producing areas, faces a rapid decline of saffron industry. Among many other factors responsible for decline of saffron industry the preponderance of erratic rainfalls and drought-like situation have become major challenges imposed by climate change. Saffron has a limited coverage area as it is grown as a 'niche crop' and is a recognized "geographical indication," growing under a narrow microclimatic condition. As such it has become a victim of climate change effects, which has the potential of jeopardizing the livelihood of thousands of farmers and traders associated with it. The paper discusses the potential and actual impact of climate change process on saffron cultivation in Kashmir; and the biotechnological measures to address these issues.

Biotech crops: imperative for achieving the millenium development goals and sustainability of agriculture in the climate change era.

Biotechnological intervention in the development of crops has opened new vistas in agriculture. Central to the accomplishment of the Millennium Development Goals (MDGs), biotech-agriculture is essential in meeting these targets. Biotech crops have already made modest contributions toward ensuring food and nutrition security by reducing losses and increasing productivity, with less pesticide input. These crops could help address some of the major challenges in agriculture-based economies created by climate change. Projections of global climate change expect the concentration of greenhouse gases to increase, aridization of the environment to increase, temperature fluctuations to occur sharply and frequently, and spatial and temporal distribution of rainfall to be disturbed-all of which will increase abiotic stress-related challenges to crops. Countering these challenges and to meet the food requirement of the ever-increasing world population (expected to reach 9 billion by 2030) we need to (1) develop and use biotech crops for mitigating adverse climatic changes; (2) develop biotech crops resilient to adverse environmental conditions; and (3) address the issues/non-issues raised by NGO's and educate the masses about the benefits of biotech crops.

In vitro cormlet production of saffron (Crocus sativus L. Kashmirianus) and their flowering response under greenhouse.

A complete protocol for the saffron cormlet production under in vitro conditions and subsequent flowering under greenhouse conditions is described. Highest number of cormlets (70.0 ± 0.30) per corm slice (explant) could be regenerated on Murashige and Skoog (MS) half strength medium supplemented with thidiazuron (TDZ) (20 µM), Indole acetic acid (IAA) (10 µM), and sucrose (40 g/l). Maximum germination (90%) of these cormlets could be achieved on MS medium containing 6-benzyl amino purine (BAP) (20 µM) and α-naphthalene acetic acid (NAA) (15 µM). In order to increase the size of the in vitro raised cormlets, these were cultured on MS medium containing TDZ (15 µM) and IAA in the range of 1.5-30 µM. Maximum increase in cormlet size could be attained on TDZ (15 µM) + IAA (12.5 µM) + sucrose (30 g/l), and the average size of cormlets was 2.5g. In another experiment, apical vegetative buds of actively growing corms were cultured for cormlet development, and corms of size 2.5g could be developed on MS medium with NAA (15 µM), BAP (20 µM), and sucrose (30 g/l). The in vitro developed cormlets were dried under shade at 25 ± 2°C for 7 d. These were then planted in small cups containing clay loam soil and kept in green house at 20 ± 2°C. In vitro developed cormlets with mean weight 2.5 g showed maximum flowering (25%) as well as vegetative growth (55%), while only 19% cormlets of 2.0 g flowered. To our knowledge this is the first report on successful flowering from in vitro raised cormlets under greenhouse.

Combined effect of dark and wounding on regeneration potential of Houttuynia cordata Thunb. leaves.

Houttuynia cordata is one of the most potential medicinal and edible wild herb whose resources have decreased sharply due to excessive exploitation. Besides its slow agamic propagation, problems of browning and non-dedifferentiation have prevented the application of micropropagation in H. cordata. Through 4 week pre-culture in darkness and wounding after 1 week pre-culture, the browning rate of leaf explants decreased significantly and resulted in efficient regeneration (20.64 +/- 5.94 adventitious buds per explant) on the induction medium. The protocol shall facilitate conservation and commercial cultivation of the endangered species.