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Showing papers by "Amit Singh published in 2012"


Journal ArticleDOI
TL;DR: A discussion of the FDA guidance on regulatory classification of pharmaceutical cocrystals of active pharmaceutical ingredients (APIs) was held in Manesar near Delhi, India, from February 2-4, 2012 as mentioned in this paper.
Abstract: The December 2011 release of a draft United States Food and Drug Administration (FDA) guidance concerning regulatory classification of pharmaceutical cocrystals of active pharmaceutical ingredients (APIs) addressed two matters of topical interest to the crystal engineering and pharmaceutical science communities: (1) a proposed definition of cocrystals; (2) a proposed classification of pharmaceutical cocrystals as dissociable “API-excipient” molecular complexes. The Indo–U.S. Bilateral Meeting sponsored by the Indo–U.S. Science and Technology Forum titled The Evolving Role of Solid State Chemistry in Pharmaceutical Science was held in Manesar near Delhi, India, from February 2–4, 2012. A session of the meeting was devoted to discussion of the FDA guidance draft. The debate generated strong consensus on the need to define cocrystals more broadly and to classify them like salts. It was also concluded that the diversity of API crystal forms makes it difficult to classify solid forms into three categories that...

734 citations


Journal ArticleDOI
TL;DR: An insight is given about the various ɛ-PL producing strains, their screening procedures, mechanism of synthesis, characterization, and its application in the medical field.

154 citations


Journal ArticleDOI
02 Jul 2012-Analyst
TL;DR: The description of pathogen detection approaches based on immobilized phage virions as well as pure recombinant RBPs are described and specific advantages of RBP-based molecular probes are discussed.
Abstract: Rapid and specific detection of pathogenic bacteria is important for the proper treatment, containment and prevention of human, animal and plant diseases Identifying unique biological probes to achieve a high degree of specificity and minimize false positives has therefore garnered much interest in recent years Bacteriophages are obligate intracellular parasites that subvert bacterial cell resources for their own multiplication and production of disseminative new virions, which repeat the cycle by binding specifically to the host surface receptors and injecting genetic material into the bacterial cells The precision of host recognition in phages is imparted by the receptor binding proteins (RBPs) that are often located in the tail-spike or tail fiber protein assemblies of the virions Phage host recognition specificity has been traditionally exploited for bacterial typing using laborious and time consuming bacterial growth assays At the same time this feature makes phage virions or RBPs an excellent choice for the development of probes capable of selectively capturing bacteria on solid surfaces with subsequent quick and automatic detection of the binding event This review focuses on the description of pathogen detection approaches based on immobilized phage virions as well as pure recombinant RBPs Specific advantages of RBP-based molecular probes are also discussed

130 citations


Journal ArticleDOI
TL;DR: The findings suggest that WhiB4 systematically calibrates the activation of oxidative stress response in Mtb to maintain redox balance, and to modulate virulence.
Abstract: Host-generated oxidative stress is considered one of the main mechanisms constraining Mycobacterium tuberculosis (Mtb) growth. The redox-sensing mechanisms in Mtb are not completely understood. Here we show that WhiB4 responds to oxygen (O2) and nitric oxide (NO) via its 4Fe-4S cluster and controls the oxidative stress response in Mtb. The WhiB4 mutant (MtbΔwhiB4) displayed an altered redox balance and a reduced membrane potential. Microarray analysis demonstrated that MtbΔwhiB4 overexpresses the antioxidant systems including alkyl hydroperoxidase (ahpC-ahpD) and rubredoxins (rubA-rubB). DNA binding assays showed that WhiB4 [4Fe-4S] cluster is dispensable for DNA binding. However, oxidation of the apo-WhiB4 Cys thiols induced disulphide-linked oligomerization, DNA binding and transcriptional repression, whereas reduction reversed the effect. Furthermore, WhiB4 binds DNA with a preference for GC-rich sequences. Expression analysis showed that oxidative stress repressed whiB4 and induced antioxidants in Mtb, while their hyper-induction was observed in MtbΔwhiB4. MtbΔwhiB4 showed increased resistance to oxidative stress in vitro and enhanced survival inside the macrophages. Lastly, MtbΔwhiB4 displayed hypervirulence in the lungs of guinea pigs, but showed a defect in dissemination to their spleen. These findings suggest that WhiB4 systematically calibrates the activation of oxidative stress response in Mtb to maintain redox balance, and to modulate virulence.

88 citations


Proceedings ArticleDOI
01 Dec 2012
TL;DR: A watermarking algorithm in spatial domain is proposed by using the Least Significant Bit (LSB) method, which is to very good compromise between performance, robustness, computational cost and quality of embedding.
Abstract: Information technology has eased the duplication, manipulation and distribution of digital data in recent times which has resulted in the demand for safe ownership of digital images. A very crucial concern for the content owners and distributors is copyright protection and content authentication. The solution to these problems is Digital Watermarking. The watermarking is the process of embedding a signal in to other signal robustly and invisibly at the same time, the embedded signal is called watermark and the other signal is called cover or host signal. In this paper we presents a brief overview of digital image watermarking techniques in spatial and frequency domain, proposed a watermarking algorithm in spatial domain by using the Least Significant Bit (LSB) method. Every technique has some advantages and disadvantages but the LSB method is to very good compromise between performance, robustness, computational cost and quality of embedding.

61 citations


Journal ArticleDOI
01 Jan 2012
TL;DR: Phage surface clustering ultimately limits the T4 phage-immobilized surface’s ability to specifically capture its host bacteria, and this is to the authors' knowledge the largest surface capture density of E. coli reported using intact T4 bacteriophages.
Abstract: Bacteriophages offer interesting alternatives to antibodies for the specific capture and detection of pathogenic bacteria onto biosensing surfaces. Procedures for the optimal chemical immobilization of lytic bacteriophages onto surfaces are presented. More specifically, the removal of lysate contaminants from bacteriophage suspensions by size exclusion chromatography significantly increases the resultant planar surface density of immobilized bacteriophages. E. coli T4 and Salmonella enterica serovar Typhimurium P22 phage systems seem to undergo highly heterogeneous adsorption to the surface, possibly explaining the observed phage clustering at higher surface densities. The T4 phage and its E. coli host were initially employed as a model system where we discovered an optimal planar surface density of phages for best bacterial capture: 18.9 ± 0.8 phages/μm2 capturing 18.0 ± 0.3 bacteria/100 μm2. Phage surface clustering ultimately limits the T4 phage-immobilized surface’s ability to specifically capture its...

56 citations


Journal ArticleDOI
TL;DR: The results of this study showed that the targeted system improved cytotoxicity of PIK75 compared to the non-targeted system and combined therapy with ceramide augmented Pik75’s therapeutic activity.
Abstract: Ovarian cancer is a debilitating disease, which needs multi-pronged approach of targeted drug delivery and enhanced efficacy with the use of combination therapeutics. In this study, we have examined the anticancer activity of PIK75 incorporated in surface functionalized nanoemulsions for targeted delivery to SKOV-3 cells. A pro-apoptotic molecule C6-ceramide was also co-delivered to augment therapeutic efficacy. EGFR and FR functionalized nanoemulsions incorporating PIK75 and C6-ceramide were characterized for particle size, surface charge, entrapment efficiency and morphology. Fluorescence and quantitative uptake studies were conducted in SKOV-3 cells to determine intracellular distribution. Cell viability was assessed using MTT assay while mechanism of cytotoxicity was evaluated using capsase-3/7, TUNEL and hROS assay. Cytotoxicity assay showed 57% decrease in IC50 value of PIK75 following treatment with EGFR targeted nanoemulsion and 40% decrease following treatment with FR targeted nanoemulsion. Combination therapy with PIK75 and ceramide enhanced the cytotoxicity of PIK75 compared to therapy with individual formulations. The increase in cytotoxicity was attributed to increase in cellular apoptosis and hROS activity. The results of this study showed that the targeted system improved cytotoxicity of PIK75 compared to the non-targeted system. Combination therapy with ceramide augmented PIK75’s therapeutic activity.

36 citations


Journal ArticleDOI
06 Jun 2012-Analyst
TL;DR: This protocol shows no sensitivity to other environmentally relevant metal ions, confirming further that change in the optical properties of gold nanotriangles in the presence of reduced mercuric ions is solely due to the strong amalgamation tendency of mercury metal.
Abstract: Mercury is a serious environmental pollutant known to have detrimental health effects in all life forms. Here, we report the use of biologically synthesized aqueous gold nanotriangles for sensitive and selective optical detection of femto-molar levels of mercury ions by exploiting the high amalgamation tendency of mercury metal towards gold. Aqueous chloroaurate ions were reduced using lemongrass (Cymbopogon flexuosus) leaf extract at room temperature to form gold nanotriangles. Mercuric (Hg2+) ions were reduced in the presence of these triangles to facilitate amalgamation and the optical properties were monitored. We observe a significant change in the longitudinal plasmon absorption band of the nanotriangles even at femto-molar concentrations of mercuric ions. High-resolution transmission electron microscopy confirms changes in particle morphology at such low concentrations. This protocol shows no sensitivity to other environmentally relevant metal ions, including Pb2+, Zn2+, Cd2+, Fe2+, Ni2+, Sr2+, Ca2+, Mn2+, and Cu2+, confirming further that change in the optical properties of gold nanotriangles in the presence of reduced mercuric ions is solely due to the strong amalgamation tendency of mercury metal.

27 citations


Journal ArticleDOI
TL;DR: Taxifolin was found to act as an inhibitor of chaperoning process and may play a potential role in the cancer chemotherapeutics.
Abstract: Hsp90 (heat shock protein 90), a molecular chaperone, stabilizes more than 200 mutated and over expressed oncogenic proteins in cancer development. Cdc37 (cell division cycle protein 37), a co-chaperone of Hsp90, has been found to facilitate the maturation of protein kinases by acting as an adaptor and load these kinases onto the Hsp90 complex. Taxifolin (a natural phytochemical) was found to bind at ATP-binding site of Hsp90 and stabilized the inactive "open" or "lid-up" conformation as evidenced by molecular dynamic simulation. Furthermore, taxifolin was found to bind to interface of Hsp90 and Cdc37 complex and disrupt the interaction of residues of both proteins which were essential for the formation of active super-chaperone complex. Thus, taxifolin was found to act as an inhibitor of chaperoning process and may play a potential role in the cancer chemotherapeutics.

23 citations


Journal ArticleDOI
TL;DR: An unusual case of giant dermoid cyst of the floor of the mouth in a 17-year female who presented with progressively increasing swelling below her tongue is reported.
Abstract: The dermoid cysts of the mouth are most frequently located on the median line of the mouth floor and are most likely caused by the retention of the germinal epithelium during the growth of the mandible and hyoid branchial arches. We report an unusual case of giant dermoid cyst of the floor of the mouth in a 17-year female who presented with progressively increasing swelling below her tongue and reviewed the relevant literature.

23 citations


Journal ArticleDOI
TL;DR: Fractionated photothermal therapy for cancer represents a new therapeutic paradigm enabled by the application of novel functional nanomaterials and dual dye-loaded mesoporous NPs retained both their NIR absorbing and NIR fluorescent capabilities after photoactivation.
Abstract: Purpose Photothermal therapy is an emerging cancer treatment paradigm which involves highly localized heating and killing of tumor cells, due to the presence of nanomaterials that can strongly absorb near-infrared (NIR) light. In addition to having deep penetration depths in tissue, NIR light is innocuous to normal cells. Little is known currently about the fate of nanomaterials post photothermal ablation and the implications thereof. The purpose of this investigation was to define the intratumoral fate of nanoparticles (NPs) after photothermal therapy in vivo and characterize the use of novel multidye theranostic NPs (MDT-NPs) for fractionated photothermal antitumor therapy. Methods The photothermal and fluorescent properties of MDT-NPs were first characterized. To investigate the fate of nanomaterials following photothermal ablation in vivo, novel MDT-NPs and a murine mammary tumor model were used. Intratumoral injection of MDT-NPs and real-time fluorescence imaging before and after fractionated photothermal therapy was performed to study the intratumoral fate of MDT-NPs. Gross tumor and histological changes were made comparing MDT-NP treated and control tumor-bearing mice. Results The dual dye-loaded mesoporous NPs (ie, MDT-NPs; circa 100 nm) retained both their NIR absorbing and NIR fluorescent capabilities after photoactivation. In vivo MDT-NPs remained localized in the intratumoral position after photothermal ablation. With fractionated photothermal therapy, there was significant treatment effect observed macroscopically (P = 0.026) in experimental tumor-bearing mice compared to control treated tumor-bearing mice. Conclusion Fractionated photothermal therapy for cancer represents a new therapeutic paradigm enabled by the application of novel functional nanomaterials. MDT-NPs may advance clinical treatment of cancer by enabling fractionated real-time image guided photothermal therapy.

Journal ArticleDOI
TL;DR: Azadirachta indica seemed to be safe and effective in colitis by its predominant effect on promoting antioxidant status and decreasing intestinal bacterial load, free radicals and myeloperoxidase responsible for tissue damage and delayed healing.

01 Jan 2012
TL;DR: A review of phytochemicals present in orchids and their medicinal properties is presented in this paper, which deals with the phytochemical properties of orchides and their properties as a backbone of traditional herbal medicines.
Abstract: From the primitive period medicinal plants have occupied a distinct place in human’s life. They have been the backbone of traditional herbal medicines and have been extensively studied because of their pharmacological importance. Orchids are one of the largest groups of Angiosperms belonging to the family Orchidaceae. A number of constituents obtained from different parts of orchid suggest biological activity. Alkaloids are nitrogenous organic heterocyclic molecules that have pharmacological effects on humans and other animals..In orchids, 214 species in 64 genera contain 0.1% or more alkaloids. Besides alkaloids, they also possess flavanoids, phenanthrenes, terpenoids, bibenzyl derivatives and other biologically active compounds. The present review deals with the phytochemicals present in orchids and their medicinal properties. © 2012 Elixir All rights reserved. Applied Botany

Journal ArticleDOI
TL;DR: It was found that core tablets containing superdisintegrant failed to produce burst drug release pattern while effervescent agent was able to do so and this approach can provide a useful means for timed release of losartan and is helpful for patients with morning surge.
Abstract: In majority of individuals blood pressure rises in the early morning hours, which lead to serious cardiovascular complications. Formulation of pulsatile system makes it possible to deliver drug at definite period of time when symptoms of the disease condition are most critical. The purpose of the present work was to develop pulsatile release tablet of losartan potassium for chronotherapy in hypertension. The prepared system consisted of a core tablet coated with versatile and safe hydrophilic cellulosic ethers such as, hydroxypropyl methylcellulose, hydroxypropyl cellulose and sodium carboxy methylcellulose to produce burst release after predetermined lag time. Various formulation factors were studied through series of test and in vitro dissolution study. It was found that core tablets containing superdisintegrant failed to produce burst drug release pattern while effervescent agent was able to do so. Results also reveal that coating composition and coating level affects lag time. Formulation containing effervescent agent in core and coated with 200 mg hydroxypropyl cellulose provide lag time of 4.5 h with 73% drug release in 6 h that followed a sigmoidal release pattern. These values were close to the desired objective of producing lag time of 5â??6 h followed by fast drug release. This approach can thus provide a useful means for timed release of losartan and is helpful for patients with morning surge.

Journal ArticleDOI

Proceedings ArticleDOI
01 Dec 2012
TL;DR: A brief overview of digital image watermarking techniques in spatial and frequency domain, advantages of frequency domain over spatial domain techniques and proposed a water marking algorithm in frequency domain by using the discrete wavelet transform are presented.
Abstract: New opportunities have been explored in social, business, entertainment and scientific fields due to the development of high speed computer networks and that of, internet in particular. Ironically, the cause for the development is apprehensive because of the use of digital formatted data. Digital media has several advantages over analog media such as high fidelity copying easy editing and high quality. Software products which hide information within digital audio, images and video files have been introduced to address these growing concerns. One of the techniques of data hiding is digital watermarking. The watermarking is the process of embedding a signal in to other signal robustly and invisibly at the same time, the embedded signal is called watermark and the other signal is called cover or host signal. In this paper we are presents a brief overview of digital image watermarking techniques in spatial and frequency domain, advantages of frequency domain over spatial domain techniques and proposed a watermarking algorithm in frequency domain by using the discrete wavelet transform.

Journal ArticleDOI
TL;DR: In this article, a bilayer resist consisting of PMMA 495/LOR 3A allowed high fabrication yields for resonators of widths ranging from 120 to 300 nm, thickness of 40 and 70 nm, and a length of 14 μm.
Abstract: Nanoimprint lithography was used to fabricate arrays of SiCN nanoscale resonators for biological analysis applications. A bilayer resist consisting of PMMA 495/LOR 3A allowed high fabrication yields for resonators of widths ranging from 120 to 300 nm, thicknesses of 40 and 70 nm, and a length of 14 μm. To our knowledge, these 120 nm resonators are the narrowest suspended structures ever fabricated via nanoimprinting. Device to device uniformity of resonant frequency was dictated by the uniformity of the tensile stress over the device layer. The 40 nm thick and 70 nm thick devices showed average resonant frequencies of 16.6±2 MHz, and 21.7±0.3 MHz, respectively. These devices were successfully employed as elements of arrays for the detection of a biological analyte. The biotin-streptavidin system was used for such purpose. The specific capture of streptavidin induced downward frequency shifts ranging from 100 to 300 kHz, corresponding to capture densities of roughly 1 to 5 molecules per 100 nm2. Negative control experiments showed no net downward frequency shifts, demonstrating the specificity of the detection.

01 Jan 2012
TL;DR: The results suggest that therapeutic inhibition of VEGFR-2 by taxifolin as a type I inhibitor may be a promising ways to retard signaling cascade of specific proteins which play crucial role in cancer proliferation and also in development of second generation type II inhibitors.
Abstract: The VEGFR-2 kinase specific intracellular signalling cascades leading to proliferation, migration, survival of endothelial cells and increased permeability of vessels which contributes to angiogenesis. ATP is essentially required by VEGFR-2 to perform phosphorylation of specific proteins and to maintain cascade downstream. Taxifolin (plant polyphenol) inhibit the VEGFR-2 kinase by binding at ATP-binding pocket revealed by molecular docking study. Further, stability of VEGFR-2 kinase-taxifolin complex is validated by molecular dynamic simulation. RMSD analysis for 3800 ps confirmed the stability of complex. Furthermore, thermodynamic stability was evidenced by stable total energy, potential energy, and, temperature and pressure profile. After MD simulation taxifolin was found to stably interact with pocket residues Cys 917 and Lys 1053 along with water molecules. These results suggest that therapeutic inhibition of VEGFR-2 by taxifolin as a type I inhibitor may be a promising ways to retard signaling cascade of specific proteins which play crucial role in cancer proliferation and also in development of second generation type II inhibitors.

Journal ArticleDOI
TL;DR: In this article, the authors presented an analysis of the volume expansion data for periclase (MgO), lime (CaO), corundum (Al2O3), spinel and spinel from 300K up to 3000K.
Abstract: We have presented an analysis of the volume expansion data for periclase (MgO), lime (CaO), corundum (Al2O3) and spinel (MgAl2O4) determined experimentally by Fiquet et al (1999) from 300K up to 3000K. The thermal equation of state due to Suzuki et al (1979) and Shanker et al (1997) are used to study the relationships between thermal pressure and volume expansion for the entire range of temperatures starting from room temperature up to the melting temperatures of the solids under study. Comparison of the results obtained in the present study with the corresponding experimental data reveal that the thermal pressure changes with temperature almost linearly up to quite high temperatures. At extremely high temperatures close to the melting temperatures thermal pressure deviates significantly from linearity. This prediction is consistent with other recent investigations. A quantitative analysis based on the theory of anharmonic effects has been presented to account for the nonlinear variation of the thermal pressure at high temperatures.

01 Jan 2012
TL;DR: Results suggest that absence of hydroxyl group substitution at C3 increases the potency of flavonoid inhibitors for cytochrome b5 reductase.
Abstract: NADH-cytochrome b5 reductase, a flavoprotein, plays a central role in many diverse metabolic reactions. NADH-cytochrome b5 reductase has been shown to be responsible for the generation of free radicals from heterocyclic amines. Flavonoids compounds share remarkable similarity in structure but showed differences in their cytochrome b5 reductase inhibition pattern. Our molecular dynamics simulation studies revealed that the difference in substitution at C3 position of ring C may lead to difference in interaction with enzyme. Absence of hydroxyl group substitution at C3 in luteolin facilitates the strong cation-π interaction between Lys185 and ring A, and C and π-π between Phe92 and ring A, and C along with h-bonding between Lys185 and oxo group. Ring B of luteolin showed strong π-π interaction with FAD. These interactions were found absent in quercetin and taxifolin. These results suggest that absence of hydroxyl group substitution at C3 increases the potency of flavonoid inhibitors for cytochrome b5 reductase.


Journal Article
TL;DR: Molecular dynamic simulation revealed that binding of taxifolin was stable at GTP binding site of different Ras forms and may lead to improper functioning of Ras in cancer cells for cancer chemotherapeutics.
Abstract: Ras proteins, the inner plasma membrane localized small G proteins, are involved in the transduction of external stimuli to its main effector Raf kinase. Point mutation in the H-Ras p21 (G12V) leads to loss of intrinsic GTPase activity so that Ras-GTP complex continuously relay signal which is associated with human cancers. Activation of oncogenic receptor tyrosine kinases also a prominent cause of continuous signal transduction through wild Ras. Taxifolin, a plant originated polyphenol, is a principal active component of several plants such as Larix gmelini. Molecular docking revealed that taxifolin captured GTP binding site in apo Ras (wild/mutant). This interaction might be valuable to target newly synthesized nucleotide unbound Ras. Molecular dynamic simulation revealed that binding of taxifolin was stable at GTP binding site of different Ras forms and may lead to improper functioning of Ras in cancer cells for cancer chemotherapeutics.

Journal ArticleDOI
TL;DR: The microbiota of the acid mine drainage was inhabited by only one or two naturally selected microbial groups extensive genome assembly was possible with their accurate taxonomical assignments, and environmental methanotroph assembled was representing the dominant population responsible for oxidation of the methane produced after defrosting of the ice.
Abstract: Next generation sequencing (NGS) has enabled us to understand the extraordinary plasticity of prokaryotic genomes. Metagenome (qualitative and quantitative analysis of eDNA) beholds the great potential of unveiling the variable regions of the individual genotypes. At the species level genomes are comprised mainly of two types of genetic content, the core genome (present in all strains) and variable genome (present only in few strains). These strain specific gene inventories are commonly known as Genomic islands (GIs) and when they are called upon using metagenomic reads they are called as Metagenomic Islands (MIs). MIs can be further explained as habitat independent gene acquisitions of a parent genotype that have been already reported from various environments. Till date, various studies have been published which can clearly explain the potential of metagenomics to not only reveal the diversity and functional gene content of previously unknown microbial diversity but also the genome recruitment of any of the enriched species or genotype [4]. Preliminary taxonomical analysis of EGTs (environmental gene tags) against any of the public sequence database can reveal the relative abundance of the actual microbial diversity (at various ranks) present at that particular environment. After quantifying the genomic fragments of dominant species, metagenomic recruitment plots can be generated using whole metagenomic data. Additionally, individual reads can be binned at various percentage identities using already sequenced neighbour as the reference genome. De novo assembly followed by assembly validation can further help to taxonomically identify the assembled genotype. Here we are sharing (ahead of publication) a metagenomic recruitment plot of Sphingobium chlorophenolicum (Fig. 1) generated by aligning metagenomics reads of a HCH contaminated dumpsite soil (un-published data Sangwan and Lal et al.). Although sequencing depth is lower, one can easily observe the genomic fragments that are completely absent or misrepresented in the following metagenomic dataset. MIs were indentified and checked for there functional status. Fig. 1 Sphingobium chlorophenolicum genome and metagenomic islands. Metagenomic reads from a HCH (Hexachlorocyclohexane) contaminated dumpsite soil were aligned on the reference genome along with the coverage parameter. Each spot on the graph represents a metagenomic ... Metagenomic reads can be reference assembled directly or contigs/scaffolds from de novo assembly of binned EGTs can be reference mapped to the present reference genome sequences. Several strategies have been used to assemble, validate and taxonomically characterizing the contigs/scaffolds obtained from de novo or reference based metagenome assemblies [1]. In acid mine drainage (AMD) analysis Tyson et al. used a whole genome assembler algorithm to assemble the long, capillary sequencing reads. Later, scaffolds were assigned to microbial groups using their average GC% percentage [1]. Assembly was validated using mate pair constraints and taxonomy was decided using partial 16S rRNA sequence analysis. As the microbiota of the acid mine drainage was inhabited by only one or two naturally selected microbial groups extensive genome assembly was possible with their accurate taxonomical assignments. Where as in Permafrost analysis [5] a draft genome assembly of a previously unknown methanotroph was obtained using De-Brujin based algorithm and single copy (total 19 genes) gene based validation. Total metagenomic data (pre-processed) was assembled on 41 K-mer length and contigs were taxonomically characterized (at various ranks) using MEGAN [2]. Coverage for individual contigs was calculated via remapping the metagenomic reads on contigs. It was observed that environmental methanotroph assembled was representing the dominant population responsible for oxidation of the methane produced after defrosting of the ice. It was the first successful attempt to assemble a draft genome from a complex matrix (soil). The most discussed and still un-standardized parameter in the target metagenome assembly is the coverage cut-off. In acid mine drainage study scaffolds with 3× to 10× coverage were assembled and assigned to various microbial groups. Where as in permafrost analysis average contig coverage was >73×. Recently Luo et al. [3] have published some methodological standards for a individual genome assemblies from short metagenomic sequencing data. Metagenome assemblies were performed with simulated metagenomic reads spiked with sequence reads of a target genome. Reference genome coverage was plotted against several parameters (single base call error, N50 length and non target sequence) and it was observed that a minimum of 20× coverage is required for a robust and valid metagenome assembly, targeting a specific genotype. Similar analysis was performed on non-simulated metagenomic datasets. Since the current studies were based upon the short sequence reads (illumina) their relevance with other NGS platforms are still to be evaluated. In summary, all of the current metagenomic assembly efforts are qualitatively in agreement with the fact that assembling and validating an environmental genome from metagenomic dataset is still not a trivial task. Latest sequencing approaches like SIP (stable isotope probing) followed by whole genome sequencing and single cell genomics can really help to assemble a complete environmental genome (enriched) from a metagenomic dataset considering the coverage and sequencing artefacts.