Introduction
The human cytochrome p450 is an isoenzyme that is responsible for various catalytic activities. The cytochrome p450 aids in metabolism of steroids, caffeine, and other drugs (Guengerich, 1992; Sachse et al., 1999). Within the CYP1 family of cytochrome p450, the two subclasses are CYP1A1 and CYP1A2 which are both regulated by the aromatic hydrocarbon receptor (AhR) (Zhou et al., 2009). Both of these gene subclass enzymes can be found on chromosome 15q21.4 (Zanger and Schwab, 2013). CYP1A2 is a liver enzyme that is responsible for caffeine metabolism (Schweikl et al., 1993).
The CYP1A2 enzyme function varies depending on which single nucleotide polymorphism (SNP) is expressed on intron 1 (Sachse et al., 1999). The specific SNP allele observed for caffeine metabolism is the rs762551 (Zanger and Schwab, 2013). The change in a single nucleotide of C or A will determine the expression of the CYP1A2 enzyme and
The use of this method determines if the allele in the individual is either homozygous AA, homozygous CC, or heterozygous (Aklillu et al., 2003). The expression of a specific allele will determine the function and rate of the CYP1A2 enzyme (Sachse et al., 1999).
The first lane is used as a standard DNA to determine the base pairs of the genomic DNA sample. Lanes 1 and 2 were used to compare to the sample genomic DNA sample to determine which SNP genotypic allele is expressed. Lane 2 contained no restriction digestion enzymes. Lane 3 and 4 both contained restriction digestion enzymes, SacI and ApaI respectively. Lane 5 contained ddH2O as a negative control for the product, resulting in no expression of DNA. Lane 4 is analyzed for determination of the individual genomic allele expressed.
Common Expression of Genotypic and Allele Frequency in a Class
Figure 2 shows the results of the electrophoresis. Lanes 5 and 7 indicate the fragments obtained when the plasmids are digested with both restriction enzymes, indicating the approximate fragment size for the hlyA gene, the pK184 plasmid and the pBluescript plasmid. This is useful for identifying the recombinant DNA needed for this experiment
Using PCR and Gel Electrophoresis to Determine Genotype. In certain situations, it is necessary to identify DNA retrieved from a sample. When there is a small sample in need of identification, Polymerase Chain Reactions are used to multiply the DNA. in the sample in many identical samples.
Amplification reaction was done in a 25.0 µL reaction mixture containing 0.4 µL DNA (from DNA extraction), 5.0 µL of 10X PCR reaction buffer, 14.2 µL of sterelized dH2O, 2.0 µL of magnesium chloride (MgCl2, 25 mM), 1.0 µL nucleotide/dNTP mix (10 Mm), and 0.4 µL of 5 u/µL Taq DNA polymerase for each primer namely respectively. The components and the volume used for the amplification reactions are listed in Table 3.2. For the reaction, PCR reaction was performed in a programmable gradient-enabled thermocycler (Bio-Rad MyCycler™ Thermal Cycler).
-Reilly Philip. Is It In Your Genes. Cold Spring Harbor Laboratory Press. 2004: 223-228. Print
In this experiment, we perform a gel electrophoresis on the DNA. In this process, the enzymes were run through the gel electrophoresis to determine their relative sizes for each of them. The results of the certain DNA fragments are used in the final step, which is to construct a map of the DNA molecule. If we use different enzymes to cut DNA, then not every restriction site will be cut by all of the enzymes. The objective of this lab is to perform restriction enzyme of digesting plasmid DNA and constructing a map of plasmid from the results made from the experiment. Using this technique we understand what a DNA restriction enzyme is and how it works. In this process, the enzymes were run through the gel electrophoresis to determine their relative sizes for each of them. By following the experiment, we determined the positions of the restriction
Graham, TE. (1998) Effects of Caffeine on Metabolism, Exercise Endurance and Catecholamine Responses and Withdrawl. London
1.4 – State why and when health and safety control equipment, identified by the principles of protection, should be used relating to types, purpose and limitations of each type, the work situation, occupational use and the general work environment, in relation to:
By analyzing the frequency of crossovers for many different alleles can trace a linear map of each chromosome.
Scrutiny of caffeine and its effects has increased dramatically in the last 20 years, due in part to an increase in consumption of caffeine. In fact, coffee consumption among young adults rose to 3.2 cups per day in 2008 from 2.4 cups per day in 2005 (Rokerya 1). For instance, in a one hour period, on Richland College’s on-campus Starbucks, the author took note of how many customers arrived and purchased a cup of coffee. Between 8:00 and 9:00 AM, there were 51 customers, implying that – especially at college - many people are dependent on coffee in the mornings. However, the results from these studies are inconclusive and often somewhat contradictory – many studies (such as that by Tetsuya Ohara et al.) show that caffeine is a great boon to
Kaplan, G., D. J. Greenblatt, M.A.Kent, and M.M. Cotreau-Bibbo. 1996. Caffeine treatment and withdrawal in mice: Relationships between dosage, concentrations, locomotor activity and A1 adenosine receptor binding. Journal of Pharmacology and Experimental Therapeutics 266:1563-1573.
The simulation study showed that the additional information of CNP could increase the accuracy of predicted genotypic value, compared to using SNP information alone in an association study. The accuracy was heavily dependent on the heritability of CNP phenotypes (correlation of CNP genotype and phenotype) (Table 3). The higher accuracy of the prediction with CNP information might also result in smaller mean squared errors of prediction (Table 4)’.
National Institute of General Medical Sciences. (2010). "21st-Century Genetics." The New Genetics, p. 74-83. Retrieved from http://publications.nigms.nih.gov/thenewgenetics/chapter5.html
This represents a large superfamily of enzymes encoded by CYP genes. They are hemoproteins with varying ...
...rtant that tests are designed to address the issue. Examples of CYP2D6 assays available are the AmpliChip, the Luminex, and AutoGenomics. These assays test for common alleles in the population , therefore rare alleles can go unreported. The AmpliChip test uses about 240 probes to screen for 30 different alleles, including all of the ones mentioned above. Similarly, the Luminex array screens for 22 SNPs, representing 17 of the common alleles and the AutoGenomics assay screens for over 20 alleles (Lyon et al., 2012). When prescribing medications, doctors should be aware of an individual’s metabolism type in order to avoid toxicity or inefficient treatment. For instance, in the case of poor and ultra rapid metabolizers, codeine should not be administered. Instead, doctors should consider prescribing another opiate such as morphine, since it bypasses the CYP2D6 pathway.
...ary part in genotypes of potential interest that human geneticists breeders, as well as evolutionary geneticists are investigating. However, although we have the capability to unravel experiments that the founders of quantitative genetics would have never imagined, but their basic, un-computational machinery that they developed is most easily adaptable to the latest analyses that will be needed. We are far from ‘letting-go’ molecular biologists from the mathematical techniques/systems, because this age in respect to genomics has been forced into accepting gratitude due to the major importance of quantitative methods as opposed to the new molecular genetics. As geneticists tend to map molecular variation as well as genomic data, quantitative genetics will be moving to the front position because of its relevance in this age of rapid advancement in molecular genetics.