Vectors for moleculars cloning - Курсовая работа

National university of life and environmental sciences of ukraine Chair of molecular genetics and biosafety Term paper Vectors For The Molecular cloning done by: third year student group №2 department of ecology and biotechnology Pereguda Olga Scientific advisor Professor Starodub N. F. Kyiv 2010 Abstract Term paper on “Vectors for the moleculars cloning” consist of two sections: conclusions list and of the references. The object of research:different vectors for the moleculars cloning. The tasks of term paper: 1) Learned the vectors for the moleculars cloning 2) Consider and study vectors of molecular cloning, and functions, properties etc. The results presented in the form of conclusions at the end of term paper. Contents Key words Abstract Conditional shortenings Introducting Literature review 1. Plasmid vectors 2. Cosmids 3. Phagemids 4. Bacteriophage vectors 4.1. Filamentous phage 4.2.Double-stranded phage 5. Scope of Present Review 6. Life cycle and genetics of Lambda 6.1. Development of Lambda 7. Phage Lambda as a vector 7.1. Size Limitation for Packaging 7.2. Transfection of Recombinant Molecules 7.3. Biological Containment 8. Phage vectors 8.1.Replacement Vectors 8.2. Insertion Vectors 8.3.Storage of Lambda Stocks Conclusion Literature Key words Cosmids - an extrachromosomal circular DNA molecule that combines features of plasmids and phage; cloning limit - 35-50 kb. DNA - a long chain polymer of deoxyribonucleotides. DNA constitutes the genetic material of most known organisms and organelles, and usually is in the form of a double helix, although some viral genomes consist of a single strand of DNA, and others of a single- or a double-stranded RNA. Enzyme - a biological catalyst, usually a protein, that can speed up a chemical reaction by lowering it’s energy of activation without being used up in the reaction. Helicase - a type of enzyme that breaks hydrogen bonds between complementary base pairs of DNA, thereby causing the double strand to spit into separate single strands. Molecular cloning - is process of creating an identical copy of DNA fragments. Phage - derivatives of bacteriophage lambda; linear DNA molecules, whose region can be replaced with foreign DNA without disrupting its life cycle. Plasmid - an extrachromosomal circular DNA molecule that autonomously replicates inside the bacterial cell. Promoter - a specific DNA sequence that serves as a binding site for RNA polymerase near each gene. Replicon - a block of DNA between two adjacent replication origins. Vector - is an agent that can carry out a DNA fragment into a host cell. Conditional shortenings BAC - Bacterial Artificial Chromosome cos - cohesice end site DNA - deoxyribonucleic acid Kb - Kilobases Kbp - Kilobase pair nt - necleotides PCR - Polymarase chain reaction pUc, pBluscript - phagemid vectors RNA - ribonucleic acid Sp6, T7 - promoters Introduction Cloning - is the process of creating an identical copy of something. In Biology, it collectively refers to processes used to create copies of DNA fragments (Molecular Cloning), cells (Cell Cloning), or organisms. The term also encompases situations, whereby organisms reproduce asexually, but in common parlance refers to intentionally created copies of organisms. In 1972, Paul Berg and colleagues made the first “artificial” recombinant DNA molecule. The molecular analysis of DNA has been made possible by the cloning of DNA. The two molecules that are required for cloning are the DNA to be cloned and a cloning vector. Cloning vector - a DNA molecule that carries foreign DNA into a host cell, replicates inside a bacterial (or yeast) cell and produces many copies of itself and the foreign DNA. Types of Cloning Vectors are Plasmid, Phage, Cosmids. Molecular cloning refers to the process of making multiple molecules. Cloning is commonly used to amplify DNA fragments containing whole genes, but it can also be used to amplify any DNA sequence such as promoters, non-coding sequences and randomly fragmented DNA. It is used in a wide array of biological experiments and practical applications ranging from genetic fingerprinting to large scale protein production. Occasionally, the term cloning is misleadingly used to refer to the identification of the chromosomal location of a gene associated with a particular phenotype of interest, such as in positional cloning. In practice, localization of the gene to a chromosome or genomic region does not necessarily enable one to isolate or amplify the relevant genomic sequence. To amplify any DNA sequence in a living organism, that sequence must be linked to an origin of replication, which is a sequence of DNA capable of directing the propagation of itself and any linked sequence. However, a number of other features are needed and a variety of specialised cloning vectors (small piece of DNA into which a foreign DNA fragment can be inserted) exist that allow protein expression, tagging, single stranded RNA and DNA production and a host of other manipul