chapter 22: genomics I- analysis of DNA

cosmid

a hybrid between a plasmid vector and phage lambda; its DNA can replicate in a cell like a plasmid or be packaged into a protein coat like a phage. cosmid vectors typically can accept DNA fragments that are tens of thousands of base pairs in length.

high throughput sequencing

ability to rapidly sequence large amounts of DNA. different technological advances contribute to this.

yeast artificial chromosome

aka YAC. the first type of artificial chromosome to be made. developed in 1987, insert within YAC can be several hundred thousand to perhaps 2 million bp in length. a few hundred YACs are sufficienct to create a contig with fragments that span entire length of chromosome. similar structurally to normal chromosomes- two telomeres, a centromere, bacterial origin of replication, a yeast origin of replication, selectable markers and a site where large chromosomal DNA piece can be inserted.

contig

collection of clones that contain contiguous, overlapping pieces of chromosomal DNA. contig represents a physical map of a chromosome. cloning vectors known as BAC's and cosmids are often used in the construction of these.

in situ hybridization

in cytogenic mapping- technique widely used to cytogenetically map the locations of genes or other DNA sequences within large eukaryotic chromosomes. chromosomes are held in place/adhered to a surface (in situ).

chromosome painting

in cytogenic mapping- uses FISH to discern sites along a specific chromosome that correspond to specific probes used.

shotgun sequencing

most efficient and inexpensive way to sequence genomes- DNA fragments to be sequenced are randomly generated from larger DNA fragments.

next generation sequence technologies

multiple technologies that use the newest possible technology in sequencing genes.

human genome project

officially began 10/1/1990, 13yr effort. goals of obtaining a genetic linkage map of human genome, a physical map of the human genome, the DNA sequence of the entire human genome, develop technology for management of human genome info, to analyze genomics of model organisms, develop programs focused on understanding and addressing the ethical, legal, and social implications of the results obtained from project, and to develop technological advances in genetic methodologies.

functional genomics

once a genome sequence is known, researchers can examine at the level of many genes how the components of a genome interact to produce the traits of an organism.

locus

plural is loci. refers to the site within a genetic map where a specific gene or other DNA segment is found.

genome

refers to the total genetic composition of an organism or species. for example the nuclear genome of humans is composed of 22 different autosomes and an X chromosome and (in males) a Y chromosome. humans also have a mitochondrial genome composed of a single circular chromosome.

puyrosequencing

relies on release of pyrosphosphate.

genome sequencing projects

research endeavors with ultimate goal of determining the sequence of DNA bases of the entire genome of a given species.

microsatellites

short repetitive sequences that are abdundantly interspersed throughout a species genome and tend to vary in length among different individuals. usually contain di-, tri-, tetra-, or pentanucleotide sequences that are repeated many times in a row. most common one in humans in sequence (CA)n where n ranges from 5 to more than 50.

subcloning

small piece of DNA from the first cosmid vector containing gene B is inserted into another vector. subcloned DNA is labeled and used as a probe to screen a cosmid library- enables the researchers to identify a second clone that extends into the region closer to gene A.

linkage mapping

studies on how genetic crosses are conducted to map the relative locations of genes within a chromosome. in eukaryotes the distance between genes is computed in map units, mu.

metagenome

the collection of genes from a particular environmental sample.

genomics

the molecular analysis of the entire genome of a species. molecular dissection process applied to a complete set of chromosomes. segments analyzed in progressively smaller pieces- locations known on intact chromosomes

proteomics

the research area with the long term goal of determining the roles of all cellular proteins to produce the characteristics of a particular cell and traits of the complete organisms.

physical mapping

third approach in which DNA cloning techniques are used to determine the location of and distance between genes and other DNA regions. in a physical map the distances are computed as the number of base pairs between genes.

polymorphic

within a population, may vary from individual to individual. distances between linked molecular markers can be determined from the outcomes of crosses.

positional cloning

a strategy in which a gene is cloned based on its mapped position along a chromosome. has been successful in the cloning of many human genes, particularly those that cause genetic diseases when mutated. include genes in cystic fibrosis, huntington's, and duchenne MD.

bacterial artificial chromosomes

aka BAC's. developed from bacterial F factors. typically contain inserts up to 300,000 bp in length or larger. easier to use than YACs because DNA is inserted into circular molecule then transformed into E. coli; more commonly used than YACs for cloning of large DNA fragments.

fluorescence in situ hybridization

aka FISH. in cytogenic mapping- most common method of in situ hybridization using fluorescently labeled DNA probes. cells are treated with agents that make them swell and fixes them onto slide; sister chromatids visualized. chromosomal DNA is denatured. a single strand DNA probe that has biotin incorporated are added. fluorescently labeled avidin binds to biotin, and that fluorescent molecule binds to the probe before it can be viewed under a fluorescence microscope. this technique is often used by clinicians to detect genetic disorders.

P1 artificial chromosomes

aka PAC's. developed from P1 bacteriophage DNA. typically contain inserts up to 300,000 bp in length or larger. easier to use than YACs because DNA is inserted into circular molecule then transformed into E. coli; more commonly used than YACs for cloning of large DNA fragments.

sequencing by synthesis

aka SBS. method that involves the identification of each nucleotide immediately after its incorporation of each nucleotide immediately after its incorporation into a DNA strand by DNA polymerase.

sequence tagged site

aka STS. the amplified region when a pair of PCR primers copies a single site within a set of chromosomes. haploid DNA STS produces single band on gel, diploid produces two.

genetic map

aka chromosome map. a chart that describes the relative locations of genes or other DNA segments along a chromosome.

cytogenetic mapping

aka cytological mapping. aimed at determining the locations of specific sequences, such as gene sequences, within chromosomes that are viewed microscopically. when stained each chromosome of a given species has a characteristic banding pattern, and genes are mapped cytogenetically related to band location.

founder

an individual that lived many generations ago and is the origin of a disease causing allele.

comparative genomics

approach that uses information form genome projects to understand the genetic variation among different populations.

mapping

in genetics, refers to the experimental process of determining the relative locations of genes or other segments of DNA along individual chromosomes. three general approaches involves- cytogenetic, linkage and physical mapping strategies

molecular marker

in linkage mapping- a segment of DNA found at a specific site along a chromosome with properties that enable it to be uniquely recognized using molecular tools such as PCR and gel electrophoresis. can possibly be used in determining the approximate location of an unknown gene that causes a human disease.

chromosome walking

one method of positional cloning. genes position relative to a marker must be known from mapping studies- provides a starting point to molecularly 'walk' toward the gene of interest.

artificial chromosomes

other cloning vectors that can accommodate much larger sizes of DNA inserts. they behave like chromosomes when inside living cells .

metagenomics

the study of a complex mixture of genetic material obtained from an environmental sample. uses include human medicine, agriculture, bioremediation, biotechnology, global change, identification of viruses, and aquatic biology.