Genomics is the study of the whole DNA sequence found in an organism.
Approaches for making sense to sequence data
Genomics deals with genome sequencing from various organisms.
Comparative genomics provides a comparison of genomes from different species to understand species evolution and gene function information.
Functional genomics helps to determine the functions of genes and the interaction of their products.
Pharmacogenomics determines how a unique peoples genome affects their drug response in the process drug development process.
- This is a rapid DNA sequencing that can help to sequence a whole genome in a reasonable time limit.
- Genomes are cut into small fragments, sequenced then reassembled to form the complete genome sequence.
- The steps for shotgun sequencing include the following:
- DNA is broken down into large fragments (150Mbps).
- The large fragments are then transferred into a vector and cloned into a bacteria for replication.
- The fragments are then organized and mapped to find their order.
- Every fragment is then cut into smaller fragments which are then sequenced.
- Computer software maps overlapping sequences to establish the order of nucleotides.
Organisms that are sequenced
- Crop plants
- Model organisms
The human genome
- It has about 25000 genes.
- The average size is 3 billion base pairs.
- The number of genes varies from 2968 in chromosome to 231 genes in chromosome Y.
- 50% of the genome has small repeating sequences.
- Genes are widely scattered on chromosomes.
- In comparative genomics, computer software is used to search for similarity regions within genomes of various species.
- The following information can be derived from comparative genomics:
- The evolutionary relationships among other species.
- Common genes between species of different organisms.
- The sequences that show unique organism features.
- To show sequences with natural selection effects.
- Functional genomics tries to show DNA sequences’ functions.
- Roles of functional genomics include the following:
- The amount of variation within a genome.
- Where the expression of genes occur
- How products from different genes and genes interact
Open reading frames
- Open reading frames are sections in DNA where genes are located.
- Open reading frames in prokaryotes can easily be located.
- Open reading frames in eukaryotes are hard to locate.
- A small portion of the human genomes has genes.
Gene expression comparison using DNA microarrays
- Microarrays are used to determine the genes that are expressed at any moment in a cell.
- DNA microarrays are slides of glass with several immobilized copies of DNA single strands.
- Steps of a microarray comparison are as follows:
- mRNA molecules are isolated from cells to be compared.
- Reverse transcription is then used to change labeled cDNA molecules from the mRNA.
- The cDNA molecules are then washed over the DNA microarray.
- Lasers are then applied to activate cDNA fluorescent tags and to observe the color patterns to determine which sequence of DNA is expressed in which type of cell.
- The sequenced genome information provides a great impact on biology branches.
- Evolutionary biologists can use the genome information to study evolutionary patterns and establish relationships between organisms.
- Geneticists share probe sequences and gene maps to identify and locate genes causing diseases.
- Developmental biologists can utilize information from comparative genomics to find important developmental importance in organisms.
- Biochemists can use DNA sequence information to predict the function and structure of a protein.
hierarchal shotgun sequencing
a genome sequencing strategy in which small DNA fragments are mapped prior to DNA sequencing.
whole genome shotgun sequencing
a genome sequencing strategy in which the entire genome is isolated and sheared into small and large fragments and cloned into vectors.
double barrel shotgun sequencing
the sequencing of DNA inserts from shotgun sequencing at both ends.
Human Genome Project
a 13-year effort with US department of health to find:
1.a genetic linkage map of the human genome
2.a physical map of the human genome
3.the DNA sequence of the entire human genome
4.technology for management of the human genome
5.analyze the genomes of other model organisms
6.develop programs focused on on ethical, legal, and social implications of results from human genome project.
the ability to rapidly sequence large amounts of DNA.
Genomic DNA is broken into small 300 to 800bp fragments which are attached to beads via adaptors and emulsified in an oil/water mixture and isolated into droplets, which are amplified into many identical products.
sequencing by synthesis
a next-generation form of DNA sequencing in which the synthesis of DNA is directly monitored to deduce base sequencing.
the study of differences of genomic variation between different populations
the total DNA content of a haploid cell. Half the DNA content of a diploid cell
the study of the structure, content, and evolution of genomes. This is also encompassed analysis of the expression and function of both genes and proteins
the study of the proteome which is the complete protein content of a cell or organism at a given moment
used to dissect the cell to understand how cellular processes worked in isolation
an effort to try and figure out how all parts of the cell (and organism) work together
a field of study that extracts biological information from large data sets such as sequences, protein interactions, microarrays, etc. It also includes data visualization
National Center for Biotechnology Information. Web based database and research interfaces that allow access to the enormous volume of data generated by genome researchers.
European Molecular Biology Laboratory. Web based database and research interfaces that allow access to the enormous volume of data generated by genome researchers.
what are the three phases in a basic PCR run
In detail, what is the exponential phase?
an exact doubling of product occurs at ever cycle. this assumes a 100% reaction efficiency. exponential amplification occurs because all of the reagents are fresh and available. the kinetics of the reaction push the reaction to favor doubling of amplicon (a small piece of DNA that has been amplified by PCR)
In detail, what is the linear phase?
this is a high variability phase. as the reaction progresses, some of the reagent (like dNTPs) are consumed as a result of amplification. the reaction starts to slow down and the PCR products are no longer doubled at each cycle.
In detail, what is the plateau phase?
the reaction has stopped and no more products are made. if left long enough, the PCR begins to degrade. Each tube or reaction plateaus at different points due to the different reaction kinetics for each sample. these differences can be seen in the plateau phase. This is the end point, where traditional PCR takes its measurements.
Explain replication in three steps
1) double-stranded DNA is in antiparallel orientation.
2) the strands are separated. Primers bind to the 3′ ends along with DNA polymerases. The dNTPs are available for incorporation into the new second strands.
3) the final products are two copies of the original DNA with each older strand interwoven with newer strands.
what are the core aims of Genome Science?
1) establish an integrated web-based database and research interface
2) assemble physical and genetic maps of the genome
3) generate and order genomic and expressed gene sequences
4) identify and annotate the complete set of genes encoded with a genome
5) characterize DNA sequence diversity
6) compile atlases of gene expression
7) accumulate functional data, including biochemical and phenotypic properties of genes
8) provide the resources for comparison with other genomes.
What are the phases to whole gene sequencing?
1) preliminary (draft) sequencing phase
2) finishing phase
3) annotating phase
In detail, what is the preliminary sequencing phase?
first phase in whole gene sequencing. it sequences the DNA up to four times allowing for 95% of the gene to be identifiable.
In detail, what is the finishing phase?
second phase in whole gene sequencing. it sequences the DNA up to eight times with no gaps and has no more than 1 error in every 10,000 base pairs.
In detail, what is the annotating phase?
third phase in whole gene sequencing. a gene is annotated when it has been recognized from a large segment of the genome sequence. Something is usually known about its cellular role at this point.
How is PCR used to obtain a genomic clone?
chromosomal DNA is first purified from cells. the first primer is then added to the population of mRNAs, and reverse transcriptase is used to make a complementary DNA strand. the second primer is then added, and a single-stranded DNA molecule is amplified through many cycles of PCR. since only the DNA between the primers is amplified, PCR provides a way to obtain selectively a short stretch of chromosomal DNA in an effectively pure form.
Explain the first cycle of PCR
it consists of three steps
1) denature- kept at a temp of 95C. this separates or melts the strands of the double-stranded DNA.
2) hybridization-the temperature is dropped between 40-60C. the cooling of DNA in the presence of a large excess of the two primer DNA oligonucleotides allows these primers to hybridize to complementary sequences in the two DNA strands.
3) synthesize- the temperature is raised to 72C. here, Taq DNA polymerase and the four deoxyribonucleoside triphosphate begin to synthesize DNA via the law of complementary base pairing. This all starts at the point here the two primers bind.