BIOMAG

This is a practical experiment steps to carry out BLAST search of a given nucleotide sequence.   (You can either watch the video or go through the steps mentioned below the video).  Follow These Steps: Open NCBI home page http://www.ncbi.nlm.nih.gov/ Click on the text box which shows all databases and change it to "Nucleotide. In the search box, type name of a protein or gene whose sequence you would like to retrieve. For example: Human Insulin. Click on Home sapience at the right corner and you will get the list of all related Homo sapience.  Select whichever you want and click on it.  You will get the data in the format of Genbank. Since BLAST accepts only FASTA format, you need to change GenBank format to FASTA format by clicking FASTA. (just below the heading line).  The new page will appear where you will get the FASTA format of the nucleotide sequence.  Copy that sequence along with code present in the format. (Not the head line).  Go bac

Gene prediction is for the prediction and determining the coding and non-coding regions present in the given stretches of the sequence. For this, several gene prediction tools are used.  1) GENSCAN:  This tool can identify introns, exons, promoter sites and polyA signals. There are other gene prediction tools which do the same. It depends on what the user terms a "Probabilistic Model" of genome sequence composition and gene structure.  2) GRAIL:  It stands for Gene Recognition and Analysis Internet Link. GRAIL 1 makes use of a neural network method to recognize coding potential in a fixed length. GRAIL 1a expands on this method by considering regions immediately adjacent to coding regions.  3) FGENEH/FGENES:  FGENEH makes use of linear discriminant analysis which predicts the internal exons by looking for structural features such as donor and acceptor splices.  FGENES can be used when multiple genes are expected to in a given DNA sequences

There are several techniques which are used for the identification of a protein structure such as optical rotatory dispersion, X-ray crystallography, spectroscopy and nuclear magnetic resonance, these techniques are powerful in determining the structural features of protein but they require many hours of highly skilled and technically dedicated work environments. There are several computational techniques which allow for the biological discovery based on the protein sequences only and/or on their comparison to protein families.  There are three main methods to identify protein and its structural features.  1) Protein Identity Based on Composition:  This involves the methods and online packages such as AA Compident, AA Compsim and Propsearch.. (AA stands for Amino Acid).  AA Compident: It uses the amino acid composition of the unknown protein to identify known proteins of the same composition.  AA Compsim: Instead of using the experimentally derived amino acid composit

Structure alignments are usually specific to protein sequences and RNA sequences. They provide the information about the secondary and tertiary structure of the protein or RNA molecule which will help in aligning the sequences more accurately.  a) DALI: It is a distance matrix alignment. It is a fragment based method for constructing structural alignments based on similarity patterns between successive hexapeptides in the query sequences. It can generate pair wise or multiple alignments and identify a query sequence's structural neighbors in the Protein Data Bank. b) SSAP: stands for Sequential Structure Alignment Program. It is a based on dynamic programming method of structural alignment that uses atom-to-atom vectors in structure space as comparison points.  Phylogenetic Analysis It refers to the studying of evolutionary relationships. Final goal is to construct the evolutionary tree describing the relationships of the various taxa with respect to each other.

There are two types of Search Tools in Bioinformatics. 1) Text Based Search Tools and 2) Sequence Similarity Based Search Tools.  1) Text Based Search Tools: a) SRS = Sequence Search and Retrieval Tool. PIR and other protein sequence database searching.  b) ENTREZ = This is for the nucleotide, GenBank, protein, DDBJ and NCBI searching. It is like a collection of these databases. c) ExPASy = stands for Expert Protein Analysis System, this is for the SWISS-PROT and other searching. 2) Sequence Similarity Based Search Tools: a) FASTA: This is a DNA and Protein sequence alignment online software package. This initially observes the word pattern hits, word-word matches of a given length, and matches the potential matches before performing more time consuming search using Smith-Waterman algorithm. The size taken for a work, given by the parameter ktup, controls the sensitivity, and speed of the program. Increasing the ktup value decreases the number of background hi

            An investigator who has an aim of inferring the evolutionary, structural and functional relationships between the sequences from different species, he would analyze the similarities and differences in the nucleotide bases or amino acid sequences. The most common method used for the comparative study is Sequence Alignment which provides the mapping between the residues of two or more sequences.  In the sequence alignment, Gaps & Insertions, Global Alignment and Local alignment must be considered based on what is the purpose of the research or study. One may consider all of these.  Gaps and Insertions : These are for studying the mutations in gene sequences. here, an investigator would achieve the better correspondence between two sequences, if he introduces a gap in sequence. similarly, he would allow an insertion in other sequence. Biologically, this corresponds, introducing a new DNA into a gene of interest.  Global Alignment : For this type of al

Extraction of chromosomal DNA from eukaryotic cells is the common procedure in molecular biology experiments and forensic analysis. Chromosomes of eukaryotic cells are located within the nucleus of the cell. Each eukaryotic chromosome consists of a single DNA molecule wrapped around the histone proteins. In order to get the chromosomal DNA out of the cell following steps are used.  1) Cells are treated with detergents such SDS (Sodium Dodecyl Sulfate), Triton-X-100 or Tween in order to break open the cell. These detergents will break the plasmamembranes and nuclear membrane of the cell and cytoplasmic material & proteins are digested. (The membranes are dissolved using detergents that solubilize the phospholipids that make up these membranes).  2) Once the membranes are dissolved, with the help of centrifuge, the cell lysate is formed. This cell lysate contains the cytoplasmic material, organelles and nuclear material and chromosomes. 3) The proteins are then

1.       Spin down 50-100 ml well-grown bacteria, 3600 rpm,15min. 2.     Resuspend bacteria with 20 ml Buffer S, immediately add 100 µl ProteinaseK (10 mg/ml). Vortex to make sure no chunks. 3.       Add 2 ml of 20% SDS, mix gently by inverting. 4.       Incubate the mixture at 65 o C for 1 hr with inverting every 15 min. 5.     Add 10 ml of phenol and 10 ml of chloroform, mix thoroughly by inverting for 5 min, spin at 3600 rpm for 20 min. 6.       Transfer supernatant to a new tube, add 0.6 volume of isopropanol. Mix gently by inverting. You will see cotton-like genomic DNA. 7.       Hook out the cotton-like DNA to a 1.5 ml tube; wash with cold 70% ethanol. 8.      Dry DNA at RT, dissolve DNA in 500 µl H 2 O (50 o C or 4 o C overnight). 9.      Add 5 µl DNase-free RNase A (20 mg/ml stock) to the DNA. Incubate at 37 o C for 30 min. 10.   Add 500 µl phenol, mix, spin at 3600 rpm for 20 min. Repeat phenol extraction once if necessary. 11.   Tr

           Bioinformatics is branch of biological and biomedical sciences and it is an application for the management of the vast biological information. Technically, Bioinformatics is the combination of biological sciences and information technology. In this field, the computers, softwares and online databases are used to gather, store, analyze and integrate biological information and genetic information. These information can then be applied to gene-based drug discovery, protein engineering and development. With the help of this, an investigator can store or analyze the genetic information in understanding the human diseases  and in the identification of new potential molecular targets and drug discovery. Bioinformatics can also be applied to study the fundamental biomedical problems.  Biological Databases: Huge amount of data of nucleotides, proteins and structures can not be written in a book.. Its gonna take a lot of time and effort to write those data.. But, tech