Polymorhpisms and SNPs
From WikiGenetics
Polymorphisms are deviations from the most common (wildtype) alleles, usually in non-coding regions. They are known to be neutral, with no observed loss or gain of function. There are quite useful as molecular markers when groups are trying to find or map new genes. There are many types of sequence polymorphisms, and many ways to test for their presence.
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[edit] Single Nucleotide Polymorphisms
(Figure: Illumina GeneChip)
According to Nature: "Single Nucleotide Polymorphism", or SNP. The most common type of change in DNA (molecules inside cells that carry genetic information). Single nucleotide polymorphisms occur when a single nucleotide (building block of DNA) is replaced with another. These changes may cause disease, and may affect how a person reacts to bacteria, viruses, drugs, and other substances.
So with this in mind and setting aside contributing environmental factors, living organisms are products of their respective genomes. An organism's genome is a code that dictates gene regulatory elements and the protein encoding genes that are transcribed, essentially all that result in a particular organism. When investigating individuals with in a particular species, for the most part they are genetically identical yet express varying characteristics. Recent studies have pointed to the importance of variations of nucleotides within the deoxyribonucleic acid (DNA) double helix at specific loci within it's genome in regard an individuality. It appears, however, that certain loci are more prone to polymorphic variation than others, at least this is the case with humans. Recnet studies suggest that miototis event occur in certain parts of the genome more so than others, this has lead to the hypothesis that so called recombination hot spots, similar to the chi sequences of microbes are of importance and Single nucleotide base (SNPs, pronounced "snips") are indicators of such hot spots(Zhao et al, 2007).
The co-inheritance of SNPs with genes of interest (i.e. disease genes) is a phenomenon that allows for SNPs to be used as markers for potential disease genes. In a situation where there are more polymorphisms that would be expected from random mutations it is possible that that segment of the genome contains a gene that has some evolutionary or physiological significance. Additionally SNPs can directly result in a phenotype change. For example the genetic basis of blue vs non-blue eye color has been linked to the OCA2 gene, where three-single nucleotide polymorphisms occurring in intron 1 (Duffy et al, 2007).
Image:Figure2.gif
(Figure: 454 Life Science Sequencer.)
It is becoming increasingly feasible to associated genetic variation to phenotypes of interest due to the advancements in the technologies that detect genetic sequences. Using the mouse model, more specifically Roquin mice. These mice exhibit a phenotype that resembles a human autoimmune disease. By crossing the Roquin mice with wild-type then screening the progeny, it is possible to determine the chromosomal location of the disease allele. The co-inheritance of the SNP and a a hypothetical disease gene is depicted below. the accompanying table shows the occurrence of the SNPs in the F2 progeny, the SNPs 4 and 5 are higher in prevalence in the affected progeny than the non-affected.
Increasingly insights into human and other organisms diversity has become evermore informative with the aid of recent technological advances that possess the ability to accurately define individual genetic variances. This allows for comparisons to be made between those presenting the phenotype of interest and those whom do not. Such association studies provide information into why one individual displays different characteristics from that of others of the same species (phenotypic diversity).
Determining genetic markers that help for clone identification. Random mutation occurs even in clones and can inheritable. Because of this it is possible to identify the origin of clones, with Bacilius anthriticus as and example; A strain used in a bioterrorism attack was successfully traced back to it lab of origin. The haemoragenic pathogen E. coli O157:H7 has been characterized in a more effective manner using micro-array and pyrosequencing methods. In the studies by Manning they evaluated using a comparative genomic hybridization analysis 83 O157 genes and locate 96 SNPs. Of interest was located in the uidA gene and was a guanosine insertion that resulted in a frameshift. It was of interest based on its late emergence and additionally explains the loss of beta-glucronidase activity of the O157 strain (Manning, et al. 2008).
A distinguishing factor of sequencing by hybridization platforms is the requirement of a reference genome by which to identify genetic aberrations. This is not the case with Sequencing by synthesis methods. , which can arize from point mutation, insertion or deletions errors in DNA repair pathways; Variable number of tandem repeats (VNTR) are another polymorphism, that comprise of repeating bases where the number of bases is the defining characteristic (i.e GGGGGGG versus GGGGGGGGGGGG or GGCGGCCACTTC, as with the below depicted example); Copy number variants, where by the gene is over expressed by duplication or overactive alterations to general transcription factors (for example if the molecular mechanism for detachment from the proximal promoter region is hindered, possible by a non-synonymous SNP).
SNPs are more often then not co-inherited with inheritable traits associated with a particular phenotype. Or depending on their location can alter: control of transcription; RNA processing; RNA transport control and localization; mRNA degradation control; protein expression control; protein activity; oncogenesis; or as markers for genes of interest(Komura, 2006. Stranger, 2007. Kwan, 2008. Jianping, 2008). SNPs that are present in greater than 1% of a population are usually taken to represent an evolutionarily significant advantage (Ledford, 2008). Linkage analysis uses this co-inheritance to provide insight into the chromosomal location of a gene of interest using a phenomenon known as linkage dis-equilibrium.
Figure: A 12 residue VNTR (yellow).
[edit] Microbial genetic diversity
Prokaryotic and eukaryotic SNPs are identified using various techniques with numerous practical applications be they medical, environmental or industrial. Using a prokaryotic example, Escherichia coli O157:H7 , 96 different loci have been identified as displaying single-nucleotide polymorphisms. These 96 loci have proved valuable for reconstructing recent evolution of the E. coli O157:H7 clade, and indicate that virulence of this clade has changed during evolution (Manning et al, 2008). Additional other studies of SNPs in this clade of E. coli have been used to identify loci in which the SNPS represent non-synonmymous coding changes that have been actively selected during evolution (Zhang et al 2006).
Microbial genomes are somewhat less complex than those of higher eukaryotes like humans and with significantly shorter generations this increases the probability of mutations occurring and being inherited. Bacterial Chi sequences are a series of eight base pairs in chromosomal DNA that promote recombination; this events increase the likelihood of genetic variations arising as a result of insertions and deletions of large segments of DNA (Zhao et al, 2007).
Scientific knowledge of the extent by which microbes differ genetically has increased and will continue to do so as the rapid, high throughput sequencing technologies' resolving power increases. The practical applications of the various technologies are numerous. The increasing resistance to antibiotics by pathogens such as Mycobacterium tuberculosis and the examination of the HIV virus will be exemplify the applications of the technologies.
[edit] Mycobacterium tuberculosis
The World Health organization reports 1.6 million deaths world wide in 2005 with the causative agent being Mycobacterium tuberculosis (TB). This a slow growing mycobacterium colonizes the lungs of human hosts untill ultimately killing the host but that is not until, if untreated, the host passes the pathogen on average to 10-15 people each year. increasing resistance to antibiotic is increasing in incidence and hindering resolution of this global problem.Mycobacterium tuberculosis with alterations within the operon encoding the arabinosyl transferase encoding gene (EmbCAB operon); particularly solitary nucleotide substitutions in codon 306 of EmbB are Ethambutol (EMB) resistant (Alcaide et al 1997, Telenti et al 1997). Pyrosequencing analysis of 28 clinical isolates rapidly identified embB306 mutations, detecting three different single-base substitutions:ATG to: ATA, ATC, ATT , GTG, CTG, ACG; leading to either Met to Val or Ile(. Mutated embB alleles were detected in 2 multidrug-resistant (MDR) EMB-susceptible strains (Sola et al 2005). This studie shows Pyrosequencing to be an effective, high though-put screen for EMB resistant strains of Mycobacterium tuberculosis.
[edit] HIV
The Human Immunodeficiency Virus (HIV) has a high capability for variation in regard to surface proteins; it is therefore quite difficult to design a widely effective HIV treatment strategy. Key in the resolution of this are toll-like receptors (TLRs) of the human hosts, which have been linked with either rapid or slow progression of infection. In particular two SNPs have been identified in humans that are associated with phenotypes in which infection progresses rapidly (Bochuda et al 2007).
The role of TLRs in HIV-1 infection is supported by studies of the interaction between HIV-1 and the innate immune cells. In particular the response of TLRs to HIV infection could increase the understanding of their interaction with HIV-1. Replication of HIV-1 is dependent on NF-kB, a TLR-induced transcription factor which promotes the activation of the HIV-1 long terminal repeats (Bafica et al 2008). This may be a influential mechanism where by HIV-1 replication is either increases or decreases disease progression (Bafica et al 2008). Due to the difficulties in sequencing individuals prior to HIV-1 infection, it would be quite revealing to obtain a individuals genomic sequence before and after HIV infection; if only to assay the variations that occur after being challenged by HIV-1.
[edit] Human Genetic Diversity
Higher eukaryotes exhibit diversification at a genetic level that compared to lower eukaryotes and prokaryotes are more dynamic with pathological and physiological significant ramifications, most of which still require further investigation in order to elucidate. It is not surprising that with in this population resides a lack of conformity amongst individuals of this population. In humans, particularly Europeans, the genetic basis of blue vs non-blue eye color has been linked to the OCA2 gene, in which three-single nucleotide polymorphisms occur in intron 1. Depending on which polymorphism is present, this imparts eye color (Duffy, 2007). Additionally, more complex examples exist; as with the Human DRD4 gene (encoding the intracellular domain of a dopaminergic receptor in the human brain) shows a high degree of genetic variation that has been linked to numerous psychological conditions including: ADHD; bipolar affective disorder; smoking and age of first intercourse (Bellgrove, 2005. Guo, 2006). In particular a VNTR (Variable Number of Tandem Repeats) has been shown to exhibit a high degree of variation, that are indicative of the above mentioned psychiatric disorders. These regions are additionally G-C rich and show minimal recombination events; indicating a strong level of natural selection (Molecular Biology of the Cell (Garland Science 2008)). Haplotypes of this type may have arisen indirectly as a result of naturally selected mutations that imparting some evolutionary advantage, these advantages however are not without antagonistic ramifications and could produce off-set mutations in progeny. For example: Mutations that increase fertility in human females may result in off-set mutations increasing homosexuality in males (Hopkin, 2004). Such off-sets seem to be equilibrating or compensating for the attributes brought forth from what appears to be an evolutionary beneficial mutation. Further investigation that broadens the sample size is required to elucidate any definitive conclusions however.
[edit] Pharmacogenomics
Medical treatments at present are hindered by individualism in terms of responses to medications that are in part due to genetic variations (i.e. SNPs; VNTR) of these individuals. Drug metabolism is of the most characterized, additionally however variations also occur within loci that once expressed are drug target sites. Pharmacogenomics incorporates such variations and their contribution to medical conditions and subsequent treatments. This is accomplished by comparing the genomic sequences from individuals exhibiting responses to treatments that vary from that of "normal" patients, hopefully unveiling a genetic variant specific to the problematic population.
Bipolar affective disorder and major depressive disorder pose many challenges from a clinical perspective. For example mood stabilizers used to treat bipolar affective disorder exhibits a wide range of individual responses which may have a strong genetic component. The ability to associate an individual's response to a particular genetic variant has become feasible with the aid of high throughput, genome-wide association studies where by hundreds of thousands of polymorphisms are able to be simultaneously tested. A major variations of interest reside with in genes that encode transporters, receptors and signal transduction molecules. For example the SLC6A4 gene (a serotonin transporter) which sequencing studies have showed a repeat polymorphism in the promoter region. This repeat polymorphism has long (l) and a short (s) forms. The (l) allele has greater transcriptional activity, and therefore more mRNA is transcribed and subsequently more protein is expressed. therefore predicting a precipitation of lower levels of synaptic serotonin (Leckbend et al 2007). This information aids in the allocation of drug therapy which is based on genotype information gathered from high throughput micro array-based genome-wide association studies, whereby candidate genes and systems can be assessed and related to affective disorders
[edit] HIV/AID
The treatment of HIV/AIDS has been hindered by the variable individual responses to medications making personalized diagnosis necessary. Using a high throughput micro-array analysis in a clinical setting would aid in the protection of the blood supply, increase preventative measures curbing the spread of the virus and optimize the allocation of medications to those already infected (Blum 2005).
[edit] Psychiatric Disorders
The role of genetic variants in diagnosing psychiatric disorders has previously been unattainable for medical practitioners until the occurrence of technological advancements such as sequencing by synthesis and DNA micro arrays have come along. VNTR play a role psychiatric disorders such as schizophrenia particularly when localized in genes encoding proteins that handle neurotransmitters such as serotonin or dopamine. Being able to rapidly detect these polymorphisms will aid in the reduction misdiagnosis, which the currently due to the "trail and error" approach is inherent with. Individuals diagnosed with schizophrenia have been shown to possess two differing polymorphisms in the SERT gene that encodes a serotonin transporter. A 17-bp variable number of tandem repeat (VNTR) polymorphism in intron 2, and a 44-bp deletion/insertion polymorphism in the promoter region of the same gene (Heils et al 1996, Herken et al 2002). Studies of this nature the PCR amplification method as described by Kaiser et al. (2001), which could be superseded by the micro-array analysis. By using a rapid high-throughput approach instead to the laborious PCR amplification method the number of samples in the cohort could be increased which would ultimately increase the reliability of the results.
[edit] Tools
[edit] Diagnostic sequencing methods
DNA sequence determination is increasingly used for diagnostic purposes. The starting point of such an approach is knowledge of sequence polymorphisms (i.e. diverse haplotypes) at loci that have physiological or pathological significance. One of the more useful tools available to distinguish an individual's haplotype is a commercial genetic sequencer, which is quite suited for this application(14). Illumina have developed such a sequencer with high accuracy for detecting single base changes, while the 454 Life Sciences sequencer detects insertion/deletion polymorphisms with extreme accuracy. Using both technologies in combination (Illumina and 454 Life Sciences sequencers), with their complimentary strengths, increases the amount of sequence data produced (Ledford, 2008). Additional support may incorporate the slightly different sequencing-by-synthesis technology available from Helicos.
“I’m after as much sequence as I can get, in a very greedy way,” says Stephen Richards of Baylor College of Medicine in Houston, Texas, who is an investigator on the drosophila project.
Recent research has employed the pyrosequencing technology developed by Roche 454 Life Science to examine and rapidly identify pathologically significant polymorphic loci. Of interest, is the increasingly antibiotic resistant Mycobacterium tuberculosis(TB), or more specifically ethambutol (EMB) (dextro-2,2′-(ethylenediimino)di-1-butanol) resistant TB. EMB is used for the treatment of multidrug-resistant (MDR) TB. Pyrosequencing analysis of 28 clinical isolates rapidly detected three different single-base substitutions to EmbB306; leading to either Met to Val or Ile. Studies have shown that strains of TB with alterations to the operon encoding arabinosyl transferase (EmbCAB operon); particularly solitary nucleotide substitutions at position 306 of EmbB (EmbB306) are EMB resistant (Sola, 2005. Braak, 2004). This information allows for the anti-microbial treatments to be tailored more specifically to this strain.
[edit] Direct sequencing Technologies
[edit] Pyrosequencing
Sequencing-by-synthesis as a method of sequence determination exploits the release of a light emitting compound upon complimentary binding of nucleotides to an immobilized single strand of DNA template (100-200 bases). The sheared sample DNA (100-200) in length has an adapter molecule added to it that allows binding to capture beads. EM-PCR amplifies the template strand, there by producing beads with multiple copies of the single stranded template strand. The platform is ready to be loaded in to the sequencer where the template strands have primers added to the 3' end allowing DNA polymerase to add nucleotides in complimentary manner. When the residues are incorporated the light released is captured by a CCD camera, with the amount of light produced is proportional to the number of residues that complimentary bind to the template strand. If multiples of the nucleotide are sequential the amount of signal released is proportional to the number of bases added to the template strand. This limits the system for if the same residue is repeated 14 or so times the signal produced is difficult to distinguish from 15 or so plus repeats ( Ronaghi, 2001).
[edit] Helicos single molecule sequencing
The process works by firstly capturing billions of single stranded sample DNA molecules on to a application specifc proprietary surface. This provides a template by which DNA polymerase to catalyze the sequence specific addition of fluorescent nucleotides to nascent complimentary strands on all templates. A washing step removes all non-bound nucleotides. Incorporated residues are imaged and positions recorded. The fluorescent group is removed leaving the incorporated residue behind. The process is then repeated for each of the four bases, and synthesizes up to 25 bases in length (Helicos Systems [1] 2008).
Helicos compares their system to pyrosequencing as: "While being very similar to pyrosequencing in that sequencing is accomplished by synthesis. The technology from Helicos used chemiluminescent labeled nucleotides that emit a signal that is detectable upon complimentary binding of just a single residue. this increases the resolving power of the assay"
[edit] Sequencing by Hybridization
Artificially synthesized oglionucleotides are immobilized via in situ photolithography, onto silicon chips. Complimentary hybridization of sample DNA to the probes releases the attached fluorophore. Using a refernece genome it is possible to determine the presence of a sequence variant, yet a sequencing by synthesis, or some other sequencing technology must be employed in order to elucidate the identity of the variant. This method can be exploited in a variety of practical applications, amoungst of which one is with in the scope of this article and that being SNP identification. By designing the oglionucleotide probes to represent sequences of interest, it is possible to determine whether or not a DNA sample consists of the sequence under investigation.
[edit] EMCCD Camera
Advances in components of direct sequencing platforms are making the $1000 genome evermore plausible. The CCD (charge-coupled device) is used in capturing the signal from the sequencing by synthesis methods. The EMCCD camera amplifies the captured signal thereby increasing the reliablity of the recorded image.
[edit] Nanopore Sequencing Technology
This method exploits the disruption of an ion channel by single nucleic acids. The level of disruption is different for each base and can discriminate modification such as methylation.
[edit] Applications
[edit] Multi locus sequence typing
This method sequences internal segments from several house keeping genes (yqiL (acetylcoenzyme A acetyltransferase), aroE (shikimate dehydrogenase)) derived from various isolates, thereby defining specific alleles at a particular locus. This information is used to determine the evolutionary relatedness of the various isolates.
[edit] Resequencing
Using as a reference a genome which has already sequenced to compare with that of different individuals "resequenced' genomes. hopefully providing insight into some pathological or physiologically significant attribute. DNA micro-arrays are ideal for this application as they are dependent on a reference genome. This can be done for either the identification of novel SNPs. This is done by using a "normal" genome as a reference, then comparing genomes from phenotypes of interest inorder to isolate SNPs which are significant to the phenotype of interest and not the reference genome. Additionally, the presance of SNPs with in an individual can be determined by using a reference genome which carries the SNPs of interest and comparing sample from un-sequenced genomes.
[edit] Phylogenetic analysis
As demonstrated by the highly accurate determination of phylogeny of E. coli 0157:H7 strains have been rapidly classified into 8 clades.
[edit] PHAT (Probe Hybridization Array Typing)
Micro-array technologies like those of Illumina (above) or Affymetrix employs artificially synthesized oglionucleotides that are bound to small glass slide or silicon microchip. The technology employs the premise that if the single stranded sample DNA binds complimentary to the oglionucleotide probes on the chip at chemiluminescent compound will be released and detected.
[edit] Copy number variants
The tools which are available to detect SNPs and VNTR () can additionally be employed to identify copy number variants (CNV). CNV are genomic structural variants in which the number of copies of a particular segment are different in two or more different genomes as compared to a "reference genome". The association of theses variants to some pathological or physiological significant phenotype are currently the major source of genetic variation and some 1,237 copy number variants have been identified in humans, thus far ([2]Iafrate, 2004; Sebat, 2004). The most important factor to keep in consideration in regard to CNV if using microarray technologies, is for the capability to accurately distinguish between 3,4 and 5 copies (Cooper, et al. 2008). The limiting factor in this case are the algorithms used to develope the probes that are used in hybridization based detection of CNV. The Affy 6.0 SNP array contains probes that posses in addtion to probes of known SNPs (Hapmap), but also have so-called "copy number probes" that also represent regions of the genome where known CNV occur (McCarrol et al, 2008).
[edit] Linkage Analysis
Marking genes using SNPs employs a methodology known as linkage analysis. The linkage between single nucleotide polymorphisms and disease genes allow inference of chromosomal positioning of disease genes (Alberts, 2006). The below figure depicts a situation where by a mother with a disease phenotype mates with a non affected male. The resultant progeny, some of which carry the disease gene and SNP, while some are non-carriers of the disease gene and yet possess the SNP, other have neither the disease gene or the SNP. SNPs in this case are used to associate the locus containing the disease gene through the aid of what is colloquially known as an association study. In comparing the sequence for affect versus non affected individuals based on the presence or absence of a particular SNP which is present in the mother and not the father and relying on the assumption that there is co-inheritance of the disease gene and the genetic marker, it is possible to determine chromosomal location of the disease gene.
[edit] Pathogen strain identification
Phylogenic typing methods have previously used restriction fragment length polymorphisms (RFLPs), or micro-satellites. More recently, using SNPs based methods such as genome-wide micro-arrays, or sequencing by synthesis, have shown a degree of inaccuracy with the previous methods. The identification of pathogenic strains using micro-array technology, such as in the the case of Baccilus anthriticus, the organism more commonly known as Anthrax. Successfully traced a strain used in a bioterroisms attack to it's lab of origin using genome-wide association studie. Additionally the enteroheamoragenic strain of Escherichia coli O157:H7 clade 8, a known fatal pathogen, was successfully typed using the SNP based method yielding more accurate results than that of previous MLST method (Manning, 2008).
[edit] Tailoring of drugs in individuals
There is great potential to minimize harm that is normally associated with conventional diagnostic methods using information obtained from genome-wide association studies. At present the FDA has approved the use of cytochrome P450 Roche Ampli-chip analysis to aid doctors in personalized treatments. Cytochrome P450 belongs to a super family of CYP2 metabolizing enzymes, comprising over 50. The way the enzymes handle psycho tropic medications are influenced by individual genetic variations with in genes encoding such enzymes (i.e P450). These variations can influence the enzymes binding site or alter enzyme function in other ways. Classifying persons under treatment as either poor, moderate or extensive metabolizes based on genotype will aid in the prevention of adverse reactions and reduce the possibility of harm caused by the metabolism of such drugs (4). This method of diagnosis could precipitate in minimizing harm caused by dosage related, adverse reactions of some medication like hepatotoxicity.
[edit] Detection of cancer susceptibility
Pharmacogenomics is advantageous in treating estrogen-receptor-positive, node-negative breast cancer whereby using high throughput micro-arrays can detect the early stages of tumor development(5). This would potentially broadening treatment options and increasing life expectancy
With the molecular mechanisms of many cancers becoming defined and the decreasing cost of genome-wide sequencing the potential for SNP based methods for prognosis of cancer is great. Resequencing a individuals genome using that individuals previously sequenced genome as reference, is likely to allow for early detection of even difficult to detect cancers and become economically feasible.
[edit] Tissue typing
The use of polymorphisms in the context of Human lymphocyte antigens is more specific than the currently used ELISA methods. Hypothetically if their are single nucleotide polymorphisms that are significantly associated with a particular type of tissue, in the context of organ transplants a more specific typing method you be available.
[edit] Discussion
Back in the early 1990's the conventional Sanger based method was prime and is still in use today with practical applications extending in to the future. The ability for the conventional Sanger methodology to be applicable in project on small and large scales is the prominent reason behind this platforms continuing success. The the system is limited by it's reliance on either having to amplify the target sequence by either PCR or by transfection in to Escherichia coli. This technology is still able to deliver highly accurate sequence date at a cost of $0.50 per kilobase. Yet, when considering the size of the human genome this is economically unfeasible, for the trend at present is to produce a Human geenomic sequence for USD$1000.00. The technologies that aim to achieve this are colloquially known as "Next generation sequencers". List amoungst this dubious title are the sequencing-by-synthesis platforms like the 454 Genome Sequencers, Roche Applied Science; Basel), the Solexa technology (used in the Illumina (San Diego) Genome Analyzer), the SOLiD platform (Applied Biosystems; Foster City, CA, USA), the Polonator (Dover/Harvard) and the HeliScope Singleh Molecule Sequencer technology (Helicos; Cambridge, MA, USA).
The commercially available technologies used in the detection of physiologically or pathologically significant polymorphisms such as SNPs and VNTR, are capable of rapid high throughput analysis with extreme accuracy. The major limiting factor being cost, mostly of the reagents that are involved in the process. Additionally the type of polymorphisms under investigation will dictate experimental design; be it use of a sequencer from either Illumina, 454 Life Science, helicos or the platform from Complete Genomics; or by combining the technologies in a efficacious fashion. The combinatorial advantages have been exemplified by the Dorsophila project, whereby the researchers whom combined the 454 sequencer with the sequencing technology from Illuminia, and yielded as much sequence data as they could get their greedy hands on (15). The sequencing by hybridization technologies increasingly important in the identification of SNPs with increases in efficiency do to work flow improvements such as bar-code multiplexed sequencing. This process used the Illumina platform in addition to fragmented sample DNA being tagged with DNA bar-codes specific for a given sample. This process allows for PCR enrichment, gel purification, cluster generation and sequencing of multiple samples (Craig W, et al. 2008). Image:SNPdisc.jpeg
The recent formation of Micro-Array Quality Control (MAQC) Consortium aims to increase the intra- and inter-platform reproducibility of micro array technologies. MAQC has confirmed that careful experimental design along with attention to data analysis, the reproducibility of micro-array data is comparable across different formats and laboratories (Nature Biotechnology - 24, 1039 (2006)).
As time progresses the role of the above discussed technologies that identify SNPs and other genetic variation will become more prominent. Sequencing by synthesis techniques as well as the sequencing by hybridization platforms will only increase in cost efficiency, thereby making it economically feasible to perform larger scale sequencing projects.
[edit] References & Links
[3]Albert TJ, et al. Direct selection of human genomic loci by microarray hybridizationNature Methods - 4, 903 - 905 (2007) Published online: 14 October 2007; Corrected online: 21 October 2007 | doi:10.1038/nmeth1111 Alcaide, F., G. E. Pfyffer, and A. Telenti. 1997. Role of embB in natural and acquired resistance to ethambutol in mycobacteria. Antimicrob. Agents Chemother. 41:2270-2273.
[4]Bellgrove MA, Hawi Z, Lowe N, Kirley A, Robertson IH, Gill M. DRD4 gene variants and sustained attention in attention deficit hyperactivity disorder (ADHD): effects of associated alleles at the VNTR and -521 SNP.Am J Med Genet B Neuropsychiatr Genet. 2005 Jul 5;136(1):81-6.
Bafica A, Scanga CA, Schito M, Chaussabel D, Sher A. Influence of co-infecting pathogenson HIV expression: evidence for a role of Toll-like receptors. J Immunol 2004; 172:7229–7234.
David Benovoy, Tony Kwan and Jacek Majewski.Effect of polymorphisms within probe–target sequences on olignonucleotide microarray experiments. Nucleic Acids Res. 2008 August; 36(13): 4417–4423.
David Benovoy, Tony Kwan and Jacek Majewski.Effect of polymorphisms within probe–target sequences on olignonucleotide microarray experiments. Nucleic Acids Res. 2008 August; 36(13): 4417–4423.
Blum RA, Wylie N, England T, French C. HIV resistance testing in the USA—a model for the application of pharmacogenomics in the clinical setting. P/rarmscotfenomics. 2005;6I21:169-179.
Bochuda P, Hersbergerg M, Taffe P, Et al Polymorphisms in Toll-like receptor influence the clinical course of HIV-1infection. AIDS 2007, 21:441–446
Nicole van den Braak a, Guus Simons, et al. A new high-throughput AFLP approach for identification of new genetic polymorphism in the genome of the clonal microorganism Mycobacterium tuberculosis. Journal of Microbiological Methods 56 (2004) 49 – 62
Cui J, et al. Identification of a novel VNTR polymorphism in C6orf37 and its association with colorectal cancer risk in Chinese population. Clinica Chimica Acta 368 (2006) 155 – 159 159. J Bray, C Clarke, G Brennen, T Muncey. Should we be ‘pushing meds’? The implications of pharmacogenomics. Journal of Psychiatric and Mental Health Nursing, 2008, 15, 357–3
Ronald A. Blum. Future of molecular diagnostics and cancer
[5]M. E. Corbella and A. Puyet. Appl Environ Microbiol.Real-Time Reverse Transcription-PCR Analysis of Expression of Halobenzoate and Salicylate Catabolism-Associated Operons in Two Strains of Pseudomonas aeruginosa. 2003 April; 69(4): 2269–2275.
[6]Duffy DL, Montgomery GW, Chen W, Zhao ZZ, Le L, James MR, Hayward NK, Martin NG, Sturm RA.Am J Hum A three-single-nucleotide polymorphism haplotype in intron 1 of OCA2 explains most human eye-color variation.Genet. 2007 Feb;80(2):241-52. Epub 2006 Dec 20.
XR Ding, J Yang, DC Sun, SK Lou, SQ Wang. Whole genome expression profiling of hepatitis B virus-transfected cell line reveals the potential targets of anti-HBV drugs. The Pharmacogenomics Journal (2008) 8, 61 – 70.
[7]Guo G, Tong Y. Age at first sexual intercourse, genes, and social context: evidence from twins and the dopamine D4 receptor gene.Demography. 2006 Nov;43(4):747-69.
[8]Isola D, et al.A Pyrosequencing assay for rapid recognition of SNPs in Mycobacterium tuberculosis embB306 region Journal of Microbiological Methods. Vol. 62, no. 1, pp. 113-120. Jul 2005
Gumbo, T. Integrating pharmacokinetics, pharmacodynamics and pharmacogenomics to predict outcomes in antibacterial therapy. CURRENT OPINION IN DRUG DISCOVERY & DEVELOPMENT 11 (1): 32-42 JAN 2008.
Gunderson, K.L., Steemers, F.J., Lee, G., Mendoza, L.G. & Chee, M.S. A genome-wide scalable SNP genotyping assay using microarray technology. Nat. Genet. 37, 549–554 (2005).
Hodges E, Xuan Z, Balija V, et al. Genome-wide in situ exon capture for selective resequencing. Nat Genet. 2007 Dec;39(12):1522-7. Epub 2007 Nov 4.
Herken H, et al. Frequency of the 17-bp variable number of tandem repeat polymorphism in Turkish schizophrenic patients. Schizophrenia Research 58 (2002) 99 – 100.
Michael Hopkin. Genes could increase male homosexuality while boosting female reproduction. Nature. (2004). doi:10.1038/news041
Iafrate, A. J. et al. Nature Genet. 36, 949–951 (204).
Jianping Hua1, David W. Craig2, Marcel Brun1, et al. SNiPer-HD: improved genotype calling accuracy by an expectation-maximization algorithm for high-density SNP arrays.J Bioinfo. Vol. 23 no. 1 2007, pages 57–63 doi:10.1093/bioinformatics/btl536
Komura D, Shen F, Ishikawa S, Fitch KR, Chen W, Zhang J, Liu G, Ihara S, Nakamura H, Hurles ME, et al. Genome-wide detection of human copy number variations using high-density DNA oligonucleotide arrays. Genome Res. 2006;16:1575–1584. [PubMed]
Kwan T, Benovoy D, Dias C, Gurd S, Provencher C, Beaulieu P, Hudson TJ, Sladek R, Majewski J. Genome-wide analysis of transcript isoform variation in humans. Nat. Genet. 2008;40:225–231
Ledford H. Population genomics for fruitflies. NATURE, Vol 453. 26 June, 2008.
Leckbend SG, Bishop JR, Ellingrod VL. Pharmacogenomics in Psychiatry. Journal of Pharmacy Practice. 2007.
Manning S, Motiwala A, Springham C. Variation in virulence among clades of Escherichia coli O157:H7 associated with disease outbreaks. Proc Natl Acad Sci U S A. 2008 March 25; 105(12): 4868–4873.
Ransohoff DF. How to improve reliability and efficiency of research about molecular markers: roles of phases, guidelines, and study design. J Clin fpidem/o/. 2007:60:1205-1219.
Ronaghi, M. Pyrosequencing sheds light on DNA sequencing. Genome Res. 11 (2001), pp. 3–11.
Sebat, J. et al. Science 305, 525–528 (2004). 011-5
Stranger BE, Forrest MS, Dunning M, Ingle CE, Beazley C, Thorne N, Redon R, Bird CP, de Grassi A, Lee C, et al. Relative impact of nucleotide and copy number variation on gene expression phenotypes. Science. 2007;315:848–853. [PubMed]
Segrè AV, Murray AW, Leu JY High-Resolution Mutation Mapping Reveals Parallel Experimental Evolution in Yeast PLoS Biology Vol. 4, No. 8, e256 doi:10.1371/journal.pbio.0040256
Shendure1 J, Robi D. Mitra2, Chris Varma1 & George M. Church1. ADVANCED SEQUENCING TECHNOLOGIES:METHODS AND GOALS. Nature Reviews Genetics 5, 335-344 (May 2004) | doi:10.1038/nrg1325
Sola D, et al. A Pyrosequencing assay for rapid recognition of SNPs in Mycobacterium tuberculosis embB306 region. Journal of Microbiological Methods. Vol. 62, no. 1, pp. 113-120. Jul 2005.
[9]David Tropel1 and Jan Roelof van der Meer. Bacterial Transcriptional Regulators for Degradation Pathways of Aromatic Compounds. Microbiol Mol Biol Rev. 2004 September; 68(3): 474–500.
Telenti, A., W. J. Philipp, S. Sreevatsan, C. Bernasconi, K. E. Stockbauer, B. Wieles, J. M. Musser, and W. R. Jacobs, Jr. 1997. The emb operon, a gene cluster of Mycobacterium tuberculosis involved in resistance to ethambutol. Nat. Med. 3:567-570.
Jin Rong Zhao, Yu Jie Bai, Qing Hua Zhang, et al. Pyrosequencing-based approach for rapid detection of rifampin-resistant Mycobacterium tuberculosis Diagnostic Microbiology and Infectious Disease, Volume 51, Issue 2, February 2005, Pages 135-137
Follow this link for a detailed explanation of the technology from Illumina [10]
[11]Anatomy of DNA
[edit] Glossary
Double-strand break: A break in both strands of a DNA molecule, as distinguished from a break in just one strand.
Linkage disequilibrium: A pattern of association between two SNPs or two loci that each have multiple alleles, such that pairs of particular SNPs or alleles, one from each locus, tend to co-occur within individuals or genomes more often than would be expected if the loci are sorting independently of each other.
Recombination: The process of one double-stranded DNA molecule joining with another; specifically in the context of meiosis, the process of two homologous chromosomes exchanging large portions of their DNA (this is also called “crossing over”).
Reference Genome. ...AGTCGATCG...
SNP. ...ATTCGATCG...
Insertion. ...AG(TCGTCGTCG)nSTCG...
Deletion. ...AGATCG...
![[10]](http://www.illumina.com/media.ilmn?Title=Sequencing-By-Synthesis%20Demo&Cap=&Img=spacer.gif&PageName=illumina%20sequencing%20technology&PageURL=203&Media=1){kind=link}