Government of India

Abstract Somatic mutations in cancer genomes are caused by multiple mutational processes, each of which generates a characteristic mutational signature 1 . Here, as part of the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium 2 of the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA), we characterized mutational signatures using 84,729,690 somatic mutations from 4,645 whole-genome and 19,184 exome sequences that encompass most types of cancer. We identified 49 single-base-substitution, 11 doublet-base-substitution, 4 clustered-base-substitution and 17 small insertion-and-deletion signatures. The substantial size of our dataset, compared with previous analyses 3–15 , enabled the discovery of new signatures, the separation of overlapping signatures and the decomposition of signatures into components that may represent associated—but distinct—DNA damage, repair and/or replication mechanisms. By estimating the contribution of each signature to the mutational catalogues of individual cancer genomes, we revealed associations of signatures to exogenous or endogenous exposures, as well as to defective DNA-maintenance processes. However, many signatures are of unknown cause. This analysis provides a systematic perspective on the repertoire of mutational processes that contribute to the development of human cancer.

Abstract Cancer is driven by genetic change, and the advent of massively parallel sequencing has enabled systematic documentation of this variation at the whole-genome scale 1–3 . Here we report the integrative analysis of 2,658 whole-cancer genomes and their matching normal tissues across 38 tumour types from the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium of the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA). We describe the generation of the PCAWG resource, facilitated by international data sharing using compute clouds. On average, cancer genomes contained 4–5 driver mutations when combining coding and non-coding genomic elements; however, in around 5% of cases no drivers were identified, suggesting that cancer driver discovery is not yet complete. Chromothripsis, in which many clustered structural variants arise in a single catastrophic event, is frequently an early event in tumour evolution; in acral melanoma, for example, these events precede most somatic point mutations and affect several cancer-associated genes simultaneously. Cancers with abnormal telomere maintenance often originate from tissues with low replicative activity and show several mechanisms of preventing telomere attrition to critical levels. Common and rare germline variants affect patterns of somatic mutation, including point mutations, structural variants and somatic retrotransposition. A collection of papers from the PCAWG Consortium describes non-coding mutations that drive cancer beyond those in the TERT promoter 4 ; identifies new signatures of mutational processes that cause base substitutions, small insertions and deletions and structural variation 5,6 ; analyses timings and patterns of tumour evolution 7 ; describes the diverse transcriptional consequences of somatic mutation on splicing, expression levels, fusion genes and promoter activity 8,9 ; and evaluates a range of more-specialized features of cancer genomes 8,10–18 .

BACKGROUND: Major depression is one of the leading causes of disability worldwide, yet epidemiologic data are not available for many countries, particularly low- to middle-income countries. In this paper, we present data on the prevalence, impairment and demographic correlates of depression from 18 high and low- to middle-income countries in the World Mental Health Survey Initiative. METHODS: Major depressive episodes (MDE) as defined by the Diagnostic and Statistical Manual of Mental Disorders, fourth edition (DMS-IV) were evaluated in face-to-face interviews using the World Health Organization Composite International Diagnostic Interview (CIDI). Data from 18 countries were analyzed in this report (n = 89,037). All countries surveyed representative, population-based samples of adults. RESULTS: The average lifetime and 12-month prevalence estimates of DSM-IV MDE were 14.6% and 5.5% in the ten high-income and 11.1% and 5.9% in the eight low- to middle-income countries. The average age of onset ascertained retrospectively was 25.7 in the high-income and 24.0 in low- to middle-income countries. Functional impairment was associated with recency of MDE. The female: male ratio was about 2:1. In high-income countries, younger age was associated with higher 12-month prevalence; by contrast, in several low- to middle-income countries, older age was associated with greater likelihood of MDE. The strongest demographic correlate in high-income countries was being separated from a partner, and in low- to middle-income countries, was being divorced or widowed. CONCLUSIONS: MDE is a significant public-health concern across all regions of the world and is strongly linked to social conditions. Future research is needed to investigate the combination of demographic risk factors that are most strongly associated with MDE in the specific countries included in the WMH.

Abstract Cancer develops through a process of somatic evolution 1,2 . Sequencing data from a single biopsy represent a snapshot of this process that can reveal the timing of specific genomic aberrations and the changing influence of mutational processes 3 . Here, by whole-genome sequencing analysis of 2,658 cancers as part of the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium of the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA) 4 , we reconstruct the life history and evolution of mutational processes and driver mutation sequences of 38 types of cancer. Early oncogenesis is characterized by mutations in a constrained set of driver genes, and specific copy number gains, such as trisomy 7 in glioblastoma and isochromosome 17q in medulloblastoma. The mutational spectrum changes significantly throughout tumour evolution in 40% of samples. A nearly fourfold diversification of driver genes and increased genomic instability are features of later stages. Copy number alterations often occur in mitotic crises, and lead to simultaneous gains of chromosomal segments. Timing analyses suggest that driver mutations often precede diagnosis by many years, if not decades. Together, these results determine the evolutionary trajectories of cancer, and highlight opportunities for early cancer detection.

Abstract A key mutational process in cancer is structural variation, in which rearrangements delete, amplify or reorder genomic segments that range in size from kilobases to whole chromosomes 1–7 . Here we develop methods to group, classify and describe somatic structural variants, using data from the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium of the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA), which aggregated whole-genome sequencing data from 2,658 cancers across 38 tumour types 8 . Sixteen signatures of structural variation emerged. Deletions have a multimodal size distribution, assort unevenly across tumour types and patients, are enriched in late-replicating regions and correlate with inversions. Tandem duplications also have a multimodal size distribution, but are enriched in early-replicating regions—as are unbalanced translocations. Replication-based mechanisms of rearrangement generate varied chromosomal structures with low-level copy-number gains and frequent inverted rearrangements. One prominent structure consists of 2–7 templates copied from distinct regions of the genome strung together within one locus. Such cycles of templated insertions correlate with tandem duplications, and—in liver cancer—frequently activate the telomerase gene TERT . A wide variety of rearrangement processes are active in cancer, which generate complex configurations of the genome upon which selection can act.

BACKGROUND: Viral epidemics or pandemics of acute respiratory infections like influenza or severe acute respiratory syndrome pose a global threat. Antiviral drugs and vaccinations may be insufficient to prevent their spread. OBJECTIVES: To review the effectiveness of physical interventions to interrupt or reduce the spread of respiratory viruses. SEARCH STRATEGY: We searched The Cochrane Library, the Cochrane Central Register of Controlled Trials (CENTRAL 2010, Issue 3), which includes the Acute Respiratory Infections Group's Specialised Register, MEDLINE (1966 to October 2010), OLDMEDLINE (1950 to 1965), EMBASE (1990 to October 2010), CINAHL (1982 to October 2010), LILACS (2008 to October 2010), Indian MEDLARS (2008 to October 2010) and IMSEAR (2008 to October 2010). SELECTION CRITERIA: In this update, two review authors independently applied the inclusion criteria to all identified and retrieved articles and extracted data. We scanned 3775 titles, excluded 3560 and retrieved full papers of 215 studies, to include 66 papers of 67 studies. We included physical interventions (screening at entry ports, isolation, quarantine, social distancing, barriers, personal protection, hand hygiene) to prevent respiratory virus transmission. We included randomised controlled trials (RCTs), cohorts, case-controls, before-after and time series studies. DATA COLLECTION AND ANALYSIS: We used a standardised form to assess trial eligibility. We assessed RCTs by randomisation method, allocation generation, concealment, blinding and follow up. We assessed non-RCTs for potential confounders and classified them as low, medium and high risk of bias. MAIN RESULTS: We included 67 studies including randomised controlled trials and observational studies with a mixed risk of bias. A total number of participants is not included as the total would be made up of a heterogenous set of observations (participant people, observations on participants and countries (object of some studies)). The risk of bias for five RCTs and most cluster-RCTs was high. Observational studies were of mixed quality. Only case-control data were sufficiently homogeneous to allow meta-analysis. The highest quality cluster-RCTs suggest respiratory virus spread can be prevented by hygienic measures, such as handwashing, especially around younger children. Benefit from reduced transmission from children to household members is broadly supported also in other study designs where the potential for confounding is greater. Nine case-control studies suggested implementing transmission barriers, isolation and hygienic measures are effective at containing respiratory virus epidemics. Surgical masks or N95 respirators were the most consistent and comprehensive supportive measures. N95 respirators were non-inferior to simple surgical masks but more expensive, uncomfortable and irritating to skin. Adding virucidals or antiseptics to normal handwashing to decrease respiratory disease transmission remains uncertain. Global measures, such as screening at entry ports, led to a non-significant marginal delay in spread. There was limited evidence that social distancing was effective, especially if related to the risk of exposure. AUTHORS' CONCLUSIONS: Simple and low-cost interventions would be useful for reducing transmission of epidemic respiratory viruses. Routine long-term implementation of some measures assessed might be difficult without the threat of an epidemic.

Socioeconomic challenges continue to mount for half a billion residents of central India because of a decline in the total rainfall and a concurrent rise in the magnitude and frequency of extreme rainfall events. Alongside a weakening monsoon circulation, the locally available moisture and the frequency of moisture-laden depressions from the Bay of Bengal have also declined. Here we show that despite these negative trends, there is a threefold increase in widespread extreme rain events over central India during 1950-2015. The rise in these events is due to an increasing variability of the low-level monsoon westerlies over the Arabian Sea, driving surges of moisture supply, leading to extreme rainfall episodes across the entire central subcontinent. The homogeneity of these severe weather events and their association with the ocean temperatures underscores the potential predictability of these events by two-to-three weeks, which offers hope in mitigating their catastrophic impact on life, agriculture and property.Against the backdrop of a declining monsoon, the number of extreme rain events is on the rise over central India. Here the authors identify a threefold increase in widespread extreme rains over the region during 1950-2015, driven by an increasing variability of the low-level westerlies over the Arabian Sea.

Abstract The discovery of drivers of cancer has traditionally focused on protein-coding genes 1–4 . Here we present analyses of driver point mutations and structural variants in non-coding regions across 2,658 genomes from the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium 5 of the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA). For point mutations, we developed a statistically rigorous strategy for combining significance levels from multiple methods of driver discovery that overcomes the limitations of individual methods. For structural variants, we present two methods of driver discovery, and identify regions that are significantly affected by recurrent breakpoints and recurrent somatic juxtapositions. Our analyses confirm previously reported drivers 6,7 , raise doubts about others and identify novel candidates, including point mutations in the 5′ region of TP53 , in the 3′ untranslated regions of NFKBIZ and TOB1 , focal deletions in BRD4 and rearrangements in the loci of AKR1C genes. We show that although point mutations and structural variants that drive cancer are less frequent in non-coding genes and regulatory sequences than in protein-coding genes, additional examples of these drivers will be found as more cancer genomes become available.

Opinions on the causes of India's growth deceleration vary. World economic growth was slower in the second half of the 1990s, and that would have had some dampening effect, but India's dependence on the world economy is not large enough for this to account for the slowdown. Critics of liberalization have blamed the slowdown on the effect of trade policy reforms on domestic industry. However, the opposite view is that the slowdown is due not to the effects of reforms, but rather to the failure to implement the reforms effectively. This in turn is often attributed to India's gradualist approach to reform, which has meant a frustratingly slow pace of implementation. However, even a gradualist pace should be able to achieve significant policy changes over ten years. This paper examines India's experience with gradualist reforms from this perspective.

PURPOSE: To assess the prevalence of refractive error and related visual impairment in school-aged children in an urban population in New Delhi, India. METHODS: Random selection of geographically defined clusters was used to identify a sample of children 5 to 15 years of age. From December 2000 through March 2001, children in 22 selected clusters were enumerated through a door-to-door survey and examined at a local facility. The examination included visual acuity measurements, ocular motility evaluation, retinoscopy and autorefraction under cycloplegia, and examination of the anterior segment, media, and fundus. Myopia was defined as spherical equivalent refractive error of at least -0.50 D and hyperopia as +2.00 D or more. Children with reduced vision and a sample of those with normal vision underwent independent replicate examinations for quality assurance in four of the clusters. RESULTS: A total of 7008 children from 3426 households were enumerated, and 6447 (92.0%) examined. The prevalence of uncorrected, baseline (presenting), and best corrected visual acuity of 20/40 or worse in the better eye was 6.4%, 4.9%, and 0.81%, respectively. Refractive error was the cause in 81.7% of eyes with vision impairment, amblyopia in 4.4%, retinal disorders in 4.7%, other causes in 3.3%, and unexplained causes in the remaining 5.9%. There was an age-related shift in refractive error from hyperopia in young children (15.6% in 5-year-olds) toward myopia in older children (10.8% in 15-year-olds). Overall, hyperopia was present in 7.7% of children and myopia in 7.4%. Hyperopia was associated with female gender. Myopia was more common in children of fathers with higher levels of education. CONCLUSIONS: Reduced vision because of uncorrected refractive error is a major public health problem in urban school-aged children in India. Cost-effective strategies are needed to eliminate this easily treated cause of vision impairment.

There has been a rapidly spreading meme in U.S. pundit and academic circles since 2010 that describes China's recent diplomacy as “newly assertive.” This “new assertiveness” meme suffers from two problems. First, it underestimates the complexity of key episodes in Chinese diplomacy in 2010 and overestimates the amount of change. Second, the explanations for the new assertiveness claim suffer from unclear causal mechanisms and lack comparative rigor that would better contextualize China's diplomacy in 2010. An examination of seven cases in Chinese diplomacy at the heart of the new assertiveness meme finds that, in some instances, China's policy has not changed; in others, it is actually more moderate; and in still others, it is a predictable reaction to changed external conditions. In only one case—maritime disputes—does one see more assertive Chinese rhetoric and behavior. The speed and extent with which the newly assertive meme has emerged point to an understudied issue in international relations—namely, the role that online media and the blogosphere play in the creation of conventional wisdoms that might, in turn, constrain policy debates. The assertive China discourse may be a harbinger of this effect as a Sino-U.S. security dilemma emerges.

Magnesium-doped ZnO (ZMO) nanoparticles were synthesized through an oxalate coprecipitation method. Crystallization of ZMO upon thermal decomposition of the oxalate precursors was investigated using differential scanning calorimetry (DSC) and X-ray diffraction (XRD) techniques. XRD studies point toward a significant c-axis compression and reduced crystallite sizes for ZMO samples in contrast to undoped ZnO, which was further confirmed by HRSEM studies. X-ray photoelectron spectroscopy (XPS), UV/vis spectroscopy and photoluminescence (PL) spectroscopy were employed to establish the electronic and optical properties of these nanoparticles. (XPS) studies confirmed the substitution of Zn(2+) by Mg(2+), crystallization of MgO secondary phase, and increased Zn-O bond strengths in Mg-doped ZnO samples. Textural properties of these ZMO samples obtained at various calcination temperatures were superior in comparison to the undoped ZnO. In addition to this, ZMO samples exhibited a blue-shift in the near band edge photoluminescence (PL) emission, decrease of PL intensities and superior sunlight-induced photocatalytic decomposition of methylene blue in contrast to undoped ZnO. The most active photocatalyst 0.1-MgZnO obtained after calcination at 600 °C showed a 2-fold increase in photocatalytic activity compared to the undoped ZnO. Band gap widening, superior textural properties and efficient electron-hole separation were identified as the factors responsible for the enhanced sunlight-driven photocatalytic activities of Mg-doped ZnO nanoparticles.

Fast detection and identification of microorganisms is a challenging and significant feature from industry to medicine. Standard approaches are known to be very time-consuming and labor-intensive (e.g., culture media and biochemical tests). Conversely, screening techniques demand a quick and low-cost grouping of bacterial/fungal isolates and current analysis call for broad reports of microorganisms, involving the application of molecular techniques (e.g., 16S ribosomal RNA gene sequencing based on polymerase chain reaction). The goal of this review is to present the past and the present methods of detection and identification of microorganisms, and to discuss their advantages and their limitations.

<h2>Summary</h2><h3>Background</h3> The burden of diabetes is increasing rapidly in India but a systematic understanding of its distribution and time trends is not available for every state of India. We present a comprehensive analysis of the time trends and heterogeneity in the distribution of diabetes burden across all states of India between 1990 and 2016. <h3>Methods</h3> We analysed the prevalence and disability-adjusted life-years (DALYs) of diabetes in the states of India from 1990 to 2016 using all available data sources that could be accessed as part of the Global Burden of Diseases, Injuries, and Risk Factors Study 2016, and assessed heterogeneity across the states. The states were placed in four groups based on epidemiological transition level (ETL), defined on the basis of the ratio of DALYs from communicable diseases to those from non-communicable diseases and injuries combined, with a low ratio denoting high ETL and vice versa. We assessed the contribution of risk factors to diabetes DALYs and the relation of overweight (body-mass index 25 kg/m² or more) with diabetes prevalence. We calculated 95% uncertainty intervals (UIs) for the point estimates. <h3>Findings</h3> The number of people with diabetes in India increased from 26·0 million (95% UI 23·4–28·6) in 1990 to 65·0 million (58·7–71·1) in 2016. The prevalence of diabetes in adults aged 20 years or older in India increased from 5·5% (4·9–6·1) in 1990 to 7·7% (6·9–8·4) in 2016. The prevalence in 2016 was highest in Tamil Nadu and Kerala (high ETL) and Delhi (higher-middle ETL), followed by Punjab and Goa (high ETL) and Karnataka (higher-middle ETL). The age-standardised DALY rate for diabetes increased in India by 39·6% (32·1–46·7) from 1990 to 2016, which was the highest increase among major non-communicable diseases. The age-standardised diabetes prevalence and DALYs increased in every state, with the percentage increase among the highest in several states in the low and lower-middle ETL state groups. The most important risk factor for diabetes in India was overweight to which 36·0% (22·6–49·2) of the diabetes DALYs in 2016 could be attributed. The prevalence of overweight in adults in India increased from 9·0% (8·7–9·3) in 1990 to 20·4% (19·9–20·8) in 2016; this prevalence increased in every state of the country. For every 100 overweight adults aged 20 years or older in India, there were 38 adults (34–42) with diabetes, compared with the global average of 19 adults (17–21) in 2016. <h3>Interpretation</h3> The increase in health loss from diabetes since 1990 in India is the highest among major non-communicable diseases. With this increase observed in every state of the country, and the relative rate of increase highest in several less developed low ETL states, policy action that takes these state-level differences into account is needed urgently to control this potentially explosive public health situation. <h3>Funding</h3> Bill & Melinda Gates Foundation; and Indian Council of Medical Research, Department of Health Research, Ministry of Health and Family Welfare, Government of India.

Histamine and its receptors (H1R-H4R) play a crucial and significant role in the development of various allergic diseases. Mast cells are multifunctional bone marrow-derived tissue-dwelling cells that are the major producer of histamine in the body. H1R are expressed in many cells, including mast cells, and are involved in Type 1 hypersensitivity reactions. H2R are involved in Th1 lymphocyte cytokine production. H3R are mainly involved in blood-brain barrier function. H4R are highly expressed on mast cells where their stimulation exacerbates histamine and cytokine generation. Both H1R and H4R have important roles in the progression and modulation of histamine-mediated allergic diseases. Antihistamines that target H1R alone are not entirely effective in the treatment of acute pruritus, atopic dermatitis, allergic asthma, and other allergic diseases. However, antagonists that target H4R have shown promising effects in preclinical and clinical studies in the treatment of several allergic diseases. In the present review, we examine the accumulating evidence suggesting novel therapeutic approaches that explore both H1R and H4R as therapeutic targets for histamine-mediated allergic diseases.

Abstract Type 2 diabetes (T2D) is a heterogeneous disease that develops through diverse pathophysiological processes 1,2 and molecular mechanisms that are often specific to cell type 3,4 . Here, to characterize the genetic contribution to these processes across ancestry groups, we aggregate genome-wide association study data from 2,535,601 individuals (39.7% not of European ancestry), including 428,452 cases of T2D. We identify 1,289 independent association signals at genome-wide significance ( P &lt; 5 × 10 −8 ) that map to 611 loci, of which 145 loci are, to our knowledge, previously unreported. We define eight non-overlapping clusters of T2D signals that are characterized by distinct profiles of cardiometabolic trait associations. These clusters are differentially enriched for cell-type-specific regions of open chromatin, including pancreatic islets, adipocytes, endothelial cells and enteroendocrine cells. We build cluster-specific partitioned polygenic scores 5 in a further 279,552 individuals of diverse ancestry, including 30,288 cases of T2D, and test their association with T2D-related vascular outcomes. Cluster-specific partitioned polygenic scores are associated with coronary artery disease, peripheral artery disease and end-stage diabetic nephropathy across ancestry groups, highlighting the importance of obesity-related processes in the development of vascular outcomes. Our findings show the value of integrating multi-ancestry genome-wide association study data with single-cell epigenomics to disentangle the aetiological heterogeneity that drives the development and progression of T2D. This might offer a route to optimize global access to genetically informed diabetes care.

The radiation field is quantized by introducing four types of photons - two transverse, one longitudinal, and one scalar. The scalar photons are treated by using an indefinite metric, and it is found necessary to modify the usual supplementary condition slightly. The present theory offers a justification for the symmetrical treatment of the four components of the electromagnetic potential, recently applied by a number of authors, and proves to be very convenient in applications. The results of physical interest, however, are the same as obtained from the ordinary formulation.

The medicinal properties of plant species have made an outstanding contribution in the origin and evolution of many traditional herbal therapies. These traditional knowledge systems have started to disappear with the passage of time due to scarcity of written documents and relatively low income in these traditions. Over the past few years, however, the medicinal plants have regained a wide recognition due to an escalating faith in herbal medicine in view of its lesser side effects compared to allopathic medicine in addition the necessity of meeting the requirements of medicine for an increasing human population. Through the realization of the continuous erosion of traditional knowledge of plants used for medicine in the past and the renewed interest at the present time, a need existed to review this valuable knowledge of medicinal plants with the purpose of developing medicinal plants sectors across the different states in India. Our major objectives therefore were to explore the potential in medicinal plants resources, to understand the challenges and opportunities with the medicinal plants sector, and also to suggest recommendations based upon the present state of knowledge for the establishment and smooth functioning of the medicinal plants sector along with improving the living standards of the underprivileged communities. The review reveals that northern India harbors a rich diversity of valuable medicinal plants, and attempts are being made at different levels for sustainable utilization of this resource in order to develop the medicinal plants sector.

Abstract Transcript alterations often result from somatic changes in cancer genomes 1 . Various forms of RNA alterations have been described in cancer, including overexpression 2 , altered splicing 3 and gene fusions 4 ; however, it is difficult to attribute these to underlying genomic changes owing to heterogeneity among patients and tumour types, and the relatively small cohorts of patients for whom samples have been analysed by both transcriptome and whole-genome sequencing. Here we present, to our knowledge, the most comprehensive catalogue of cancer-associated gene alterations to date, obtained by characterizing tumour transcriptomes from 1,188 donors of the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium of the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA) 5 . Using matched whole-genome sequencing data, we associated several categories of RNA alterations with germline and somatic DNA alterations, and identified probable genetic mechanisms. Somatic copy-number alterations were the major drivers of variations in total gene and allele-specific expression. We identified 649 associations of somatic single-nucleotide variants with gene expression in cis , of which 68.4% involved associations with flanking non-coding regions of the gene. We found 1,900 splicing alterations associated with somatic mutations, including the formation of exons within introns in proximity to Alu elements. In addition, 82% of gene fusions were associated with structural variants, including 75 of a new class, termed ‘bridged’ fusions, in which a third genomic location bridges two genes. We observed transcriptomic alteration signatures that differ between cancer types and have associations with variations in DNA mutational signatures. This compendium of RNA alterations in the genomic context provides a rich resource for identifying genes and mechanisms that are functionally implicated in cancer.

The clustered, regularly interspaced, short palindromic repeats (CRISPR) and CRISPR-associated protein (Cas) system has been used as an efficient tool for genome editing. We report the application of CRISPR-Cas-mediated genome editing to wheat (Triticum aestivum), the most important food crop plant with a very large and complex genome. The mutations were targeted in the inositol oxygenase (inox) and phytoene desaturase (pds) genes using cell suspension culture of wheat and in the pds gene in leaves of Nicotiana benthamiana. The expression of chimeric guide RNAs (cgRNA) targeting single and multiple sites resulted in indel mutations in all the tested samples. The expression of Cas9 or sgRNA alone did not cause any mutation. The expression of duplex cgRNA with Cas9 targeting two sites in the same gene resulted in deletion of DNA fragment between the targeted sequences. Multiplexing the cgRNA could target two genes at one time. Target specificity analysis of cgRNA showed that mismatches at the 3' end of the target site abolished the cleavage activity completely. The mismatches at the 5' end reduced cleavage, suggesting that the off target effects can be abolished in vivo by selecting target sites with unique sequences at 3' end. This approach provides a powerful method for genome engineering in plants.