DNA methylation regulated genes in cancer:
DNA methylation is the key epigenetic change that is responsible for altered gene expression. While there is a global demethylation of human genome during tumourigenesis, selective methylation of tumour suppressor genes may also occur during this process. Oral cancers, one of the frequent disease that lead to mortality, progresses through distinct phases such as leukoplakia, erythroplatia, fibrosis etc., before culminating into sqamous cell tumours. The working hypothesis is that there is a significant alterations in DNA methylation that may lead to inactivation of tumour suppressor genes or activation of oncogenes during early stages of tumour progression. We intend to identify and characterise these critical genes as a marker for early tumour detection and possibly for therapy.

Genomics of human cancers:
Identification of potential targets that influence either restoration of normal tissue homeostasis or disruption of tumour-induced microenvironment and evaluation of these in clinical practice is envisaged. Experiments to understand this is expected to generate potential tumour progression related molecules, targets for therapy and further the understanding of tumourigenesis. An environmental influence such as chemicals and radiation on chromosomal instability is the initial focus. We are proposing to evaluate tumour cell heterogeneity not only with respect to progression but also treatment using high throughput screening by genome-wide scanning for instability (MSI), expression profiling by arrays and detection of SNPs for drug response.

Characterisation of adult human stem cells in artificial skin:
Cellular replacement therapy using adult stem cells is a promising treatment modality for several human diseases. In this context, study of skin stem cells in its natural environment is impossible. Towards this goal, we have established artificial skin reconstructs with the components of human skin. This model facilitates study of differentiation of stem cells under various extracellular cues to a given lineage and modulating the same through artificial means. This concept has applications to not only in understanding the stem cell behaviour in adult skin but also has the potential to be extended to clinical management.

Chemo-response modifications in human leukaemia:
Combination chemotherapy using mechanism based approaches has led to substantial improvement in the clinical management of human leukaemia. Traditionally used anticancer drugs suffer from innumerable side effects causes of which are minimally understood. Therefore, in lieu with the principles of pharmacogenomics, elucidation of genes regulated through the anticancer actions of several drugs is in place. Thus the identification of sequentially altered genes will lead to unravelling of new targets genes and are more amenable to transfer to the clinics.

Development of Arrays for Pathway Specific Pharmacogenomic Diagnosis of Human Diseases:
The detection and genotyping of polymorphisms and mutations has become one of the most challenging and often expensive and time-consuming obstacles to many molecular genetic applications including clinical diagnosis, pharmacogenomics and forensic analysis. Using traditional methods researchers were able to survey a relatively small number of genes at a time. The aim of this proposal is to develop and translate array based diagnosis of DNA polymorphisms in human diseases. These polymorphic genes are cumbersome to analyse one at a time and we would like to develop simple, cost effective, robust yet reliable arrays which are amenable for evaluation for pharmacogenomic purposes.

Molecular genetics of human diseases.
The study of the molecular genetic basis of human diseases to know more about the functional mechanisms of pathogenesis and its application to develop techniques for detection, confirmation of clinical diagnosis and prenatal diagnosis, is a core area of research at the Manipal Life Sciences Centre. Research on various aspects of pharmacogenomics and molecular diagnosis of several human disorders and related translational research programs are also carried out. The fully equipped automated DNA sequencing facility and accessory facilities including real-time PCR and documentation units are employed for analysis of variations in genes underlying human disorders such as sickle cell anaemia, thalassemia, haemolytic anemias and other hemoglobinopathies, Wilson’s disease, Gilbert’s disease, cystic fibrosis, fragile-X syndrome, hemochromatosis, and various cancers to mention only a few. Prenatal diagnosis for known diseases causing mutations is performed where there is a risk. . Fragment length polymorphism analysis of genomic regions with microsatellite marker loci, is performed using quantitative fluorescence PCR (QF-PCR) technique for prenatal diagnosis of chromosomal copy number variations.

Construction and Characterization of Indian Human BAC Library
The availability of genomic sequence information and complete bacterial artificial chromosome (BAC) libraries for human genome and a broad phylogenetic spectrum of organisms is now ushering in a new era in biological research and disease modeling. Bacterial Artificial Chromosomes (BACs) have become became the widely used resource for several reasons. These include large insert carrying capacity (100–300 kb), high clonal stability, low rate of chimerism, and the ease with which they can be handled. In support of the functional analysis of genes, the BACs are very useful for making transgenic animals with segments of human DNA. BACs have served as the primary source of archived genomic DNA for a variety of genome mapping, copy number studies and sequencing projects. We have constructed Indian Human BAC library. The library includes 100,224 clones. We have developed a pooling strategy and a three-step PCR screen for the Indian BAC library to isolate desired BAC clones. Screening pools will allow PCR-based screening of the Indian genome for any segment of interest. This library represents the first Indian Human BAC library and it can serve as a genetic resource facilitating medical, diagnostic, pharmacological, physiological and evolutionary studies. 

In vitro culture & in vitro mutagenesis in plants for somaclonal variation
Mutations are known to enhance the genetic variability of crop plants. Efforts are being made to improve the genetic make-up of important food crops for higher yield, oil content and development of cultivars resistant to diseases and pests. Tissue culture together with induction of mutations may be an effective way for crop improvement. Our aim is to induce somaclonal variations through in vitro mutagenesis in food crops and medicinal plants for increased yield and secondary metabolite production.

Study on Nematode infection in brinjal (Solanum melongena L.)
Brinjal (Solanum melongena L.) crop faces excessive damage, loss of fruit and decrease in yield every year. Such damage is caused by a number of abiotic and biotic factors. The major biotic factor is nematode infection, which mainly affects the root system from seedling stage. The symptoms are seen in roots (lesion) and aerial parts of plant. Therefore, we plan to investigate the mode of nematode infection, identify the genus and species and formulate a method to control the same. This study will be the platform for designing biological and chemical methods to control nematode infection. 

Genetic manipulation of plants for fungal disease resistance
A vast number of plant pathogens such as viriods that are of a few hundred nucleotide long can cause various pathogenic effects in higher plants ranging from mild symptoms to catastrophes resulting in massive damage to food crops. With more than 800 million of the world population surviving on less than a $1 a day, at least 10% of the global food production is lost to plant diseases annually, the main causative agent being fungi. An average of 12% of world crop production is lost per annum as a direct result of microbial infection. Genetic manipulation of important food crops with overexpression of disease resistance gene(s) to enhance the fungal resistance in plants is crucial.

Secondary metabolites from microfungi
The diversity of microfungi is a matter of dispute among mycologists and the consensus is that thousands of species of fungi remain unexplored especially so in the tropics. The tremendous potential of these microfungi in eliciting compounds of pharmaceutical and biological importance is well known; several secondary metabolites have been isolated and used as life saving drugs and the list is growing day-by-day. It is in this regard that we are carrying out research to explore these ubiquitous microfungi using state-of-the-art facilities available in MLSC. 

Metal Analysis in Biological Samples using AAS
The essential trace elements are of great importance for normal growth and development of the body despite their low concentrations. There are more than twenty trace elements which play a key role in biological systems and are vital for normal functioning of the human body, accompanied by some toxic metals which have no biological significance. In some cases trace elements serve as constituents of vital biological molecules and in others they are either part of enzyme system or exert their influence as co-factors for various reactions mediated by certain enzymes.  The main focus of research in our lab is on biomonitoring, with respect to exposure of various metals in an attempt to assess the potential health risk induced by these substances, for which there are no precise markers till date. We also use AAS primarily to analyse diagnostic and research samples referred to us by various hospitals and research centers.

Development of Automated Karyotyping software
Karyotype is a standardized arrangement of all the chromosomes of a cell. The chromosomes are arranged and displayed in a standard format known as an ideogram which is arranged in pairs, ordered by size and position of centromere for chromosomes of the same size. Karyotypes are used to study chromosomal aberrations. The objective of this work is to design open source software for automated karyotyping. The software takes the karyotyping image and automatically classifies chromosomes as per their length. The software also has utility to enhance the images using contrast enhancement with thresholding techniques. To identify individual chromosomes as objects we have used contour tracing algorithm (Moore’s algorithm). For statistical analysis we have implemented ImageJ package of NIH. This project is a joint project between Manipal Life Sciences Center and Manipal Center for Information Science.

Development of Post epigenomic analysis pipeline
DNA methylation is a frequent, dynamic biochemical modification in eukaryotic DNA, which is considered as crucial factor in causing cancers. It affects the C5 position of the Cytosine that belongs to CpG sites. The objective of this project is to further analyze the hyper- or hypo-methylated microarray probe sequences with respect to prediction of CpG island with principle of Gardiner and Gardiner, promoter analysis with dynamic programming concept, prediction of transcriptional factor binding sites with Hidden-Markov model algorithm. The entire project is done through Plone as a web server and Python as a scripting language.