Differentiation in Caenorhabditis Elegans - Essay Example

Differentiation in Caenorhabditis Elegans No Lecturer) Brief Introduction The starting point of the understanding of Caenorhabditiselegans was the year 1900. It is a nematode (roundworm) and one of the ideal organisms used in the understanding of biological system function. It is specifically used in the study of gene regulation and function (Kulkarni, 2008). C. Elegans was also used in this study because it is a eukaryote. This means that it has biological information that may be directly applicable in more complex-organisms such as human being. Eukaryote is small in size and it is estimated to have a genome of 9.7 X-107 base pairs or 97 Megabases. When comparing it to that of human being, it is small since humans have a large size genome (Yamamoto, 2011). Moreover, C. Elegans has a fast and suitable life cycle and can be manipulated in the laboratory. Therefore, C. Elegans was the first animal to have its genome completely sequenced. C. Elegans can be described as a simple organism that has a simple body system like excretory system that will be explained later in this report. However, due to the simplicity, its cell lineage was completely mapped for all the 959 somatic cells in the adult worm. This report will also address in brief anatomy, genetic, and the life cycle of this cell. Genetics One thing known about c. Elegans is that it has six-chromosomes five of which pairs and form autosomal. The sixth chromosome is a sex-chromosome. Hermaphrodites however, have two X-chromosomes while the male has one X-chromosome thereby naming it as hemizygous. This means that they have only one chromosome instead of a pair. The one X-chromosome can therefore be produced if the hermaphrodite has lost one of the X-chromosome. It can also happen by mating with a male (Kulkarni, 2008). Life cycle Figure 1 From the above genetic explanation, one of the reasons scientist worked with c. Elegans is because of its short life cycle as shown in Figure 1. Its life cycle depend on the surrounding temperatures. For instance, if the temperatures are 15 degree Celsius, it takes five days to complete the lifecycle. If the temperatures are increased to 20 degrees Celsius, it will take three and a half days. Now, when the temperatures are 25 degrees Celsius, it will take 2.2 days to complete the same reproductive life cycle. The egg-fertilization of C. Elegans takes place in the adult hermaphrodite that it lays for few hours. After about 40-cell stage, the eggs hatch and c. Eleganse develop through four larval stages (Kulkalmi, 2011, p. 2). When it reaches the adult stage, it produces about 300 progeny and stays alive for around two weeks. In some occasion, especially when there is food deprivation or if c. Elegans is growing in a plate at the laboratory, it can take an alternative form of life known as the dauer larval stage as shown in Figure 1. Dauer larvae cannot eat since their mouths are plugged and also it has a thin body. Remarkably, dauers can stay alive for three months. However, they seem to be non-aging because they live for months until they consume food. It is after taking food that dauers enter the L4 stage where they live for more than the normal c. Elegans. It is because of this that c. Elegans leads to further research regarding anti aging for human benefits. Promoter trapping Promoter trapping was developed before genome sequence project. It was used to screen the gene expression within an organism. In c. Elegans, a reporter bacterial gene called lacZ was involved with encoding of the enzyme ß-galactosidase. This enzyme is fused to the regulatory elements of different genes including c. Elegans that is expresses cells by ß-galactosidase (Finney M, 2012, pp. 1-2). In order to generate a promoter trapping in c. Elegans, DNA fragment need to be extracted from c. Elegans by restriction enzyme. It is then transferred to a reporter gene that has or has no promoter. Following this mixture of DNA and reporter, it induces to host organism or in this case into c. Elegans to observe the expression (Kulkarni, 2008). In this practical, both UL1 and UL6 are already fused with lacZ gene in their regulatory elements. This is used to determine the site of expression by histochemical staining. The histochemical staining is seen in this practice X-gal. This staining is colourless substrate of ß- galactosidase. When it is cleaved, it gives an insoluble blue product that is a precipitate at the sites of ß-galactosidase activity. Materials and methods This study was done in pairs such that two strains of the nematode worm Caenorhabditiselegans, UL1 and UL6 are provided for each pairs, each student worked with one stain (Kulkarni, 2008, p. 12). Both of the worms stain is washed separately by distilled water using eppendorf tube for 5 minutes. Later the worms are settled at the bottom of tube where they each are transferred to a multiwell glass or microscope slide. Each worm slide is labelled and then covered with coverslip. UL1 and UL6 slides are then placed on a metal plate on dry ice to freeze the worm and penetrate the stain. 3 minutes later, the slides are transferred quickly to an acetone at -20 degrees Celsius. Then the slide is left to dry on the bench. 3μl of X-gal staining solution is added in each well of the slide to easily determine the site of expressions later. Then the slide is put in an incubator for 1hr at 37 degrees Celsius. Afterwards, bothUL1 and UL6 stain specimens are observed under the microscope. Results C. Eleganse is found to consist of two types of sex-cells that are hermaphrodite or a male. The research shows that the hermaphrodites develop from females. This happens in both hermaphrodites and females, which are found to be somatically identical. The main difference is that hermaphroditic can produce both sperm and egg. Significantly, the predominate sex-of c. Elegans is hermaphrodites. The sperm in hermaphrodites and is produced and stored in the nematode’s genitals such that oocytes are later produced in the ovary. The egg of an adult hermaphrodite becomes fertilized when it moves to the uterus with its sperm. In self-fertilisation, all produced progeny are hermaphroditic. This means that the male c. Elegans mates with hermaphroditic as shown in Figure 2. In such a situation, the sex-ratio changes to 50 percent due to hermaphrodites and 50 percent of male (Kulkalmi, 2011, p. 14). The male sperms are found to have a competitive advantage over endogenous hermaphroditic sperm. Moreover, the male genotype is XO while Hermaphroditic is XX. Figure 2: Sex-and the single nematode Figure 3 This figure gives a closer look at the organization of the hermaphrodite gonad ß- Galactosidase expression patterns are observed under a microscope in such a way that the strains UL1 and UL6 look similar but in different sites. In UL1 hermaphrodite, the expression expresses the middle section of the hermaphrodite body. Precisely, the embryos are traced within the uterus near the vulva. A large number of hermaphrodites are found to be pregnant as their uterus is large than usual. The colour of the expression also between the green-blue and the very clear are also observed in developed embryo (Figure 3). UL6 on the other hand, has the expression in the excretory cell near the anterior of the body. This expression was found to be in green and was not that clear as much as UL1. It is not able to see the H shape in UL6 like the small round spot between figure 3 and figure 4. This has been resulted to the experiment limitation such as inquorate microscope, short time, and experiment error (Kulkalmi, 2011). Figure 5 shows the an adult hermaphrodite in different stages of development Discussion The results showed different sights of expression in UL1 and UL 6 because of inducing reporter gene in this worm (Yamamoto, 2011). When ß- galactosidase protein fuses through cell membrane, it causes enzyme inactivation. This leads to the production of signals from the organism trans-membrane to retain ß- galactosidase activity in cytoplasm resulting to an expression. In UL6, the expression detected an excretory cell (Finney M, 2012). This excretory cell comes under excretory system that is composed of four types of cells such as pore cell, duct cell, the canal cell or the large H-shaped excretory cell and the binucleate, A-shaped gland cell (Figure 5). The study shows that they have their nuclei located in the head region. Simply, this means that it is located on the ventral side of the pharynx-and pharyngeal-intestinal valve. The excretory cell contributes in waste elimination and osmotic/ionic regulation. Moreover, the shape of these cells is highly variable in nematodes. Figure 6 In UL1, the expression is observed in early embryo in the uterus of adult hermaphrodite (Kulkarni, 2008, p. 3). There is no sign of expression in neither the development embryo nor the adult or even other cells. In Hope (1991), he illustrated that the expression was found in early embryo and other type of cells such as muscle cells, nerve cells, cells of the gonad, hypodermal cells and undifferentiated cells (Kulkalmi, 2011, p. 2). However, these results are not all found in one or the same hermaphrodite but different strain of hermaphrodite (Figure 6). C. Elegans: A Practical Approach Ian A. Hope‬Figure 7 References Finney M, R. G. (2012). The C. Elegans cell lineage and differentiation gene unc-86 encodes a protein with a homeodomain and extended similarity to transcription factors. Retrieved May 12, 2014, from NCBI: http://www.ncbi.nlm.nih.gov/pubmed/2903797 Kulkalmi, M. (2011). C. Elegans as a Model System. General Biology of C. Elegans . Kulkarni, M. (2008). Role of Ubiquitination in Caenorhabditis Elegans Development and Transcription Regulation During Spermatogenesis. London: ProQuest. Yamamoto, D. (2011). Genetics of Sexual Differentiation and Sexually Dimorphic Behaviors. New York: Academic Press. Waksman Student Scholars Program. (ND). The C. Elegans Project: Introduction. Available: http://avery.rutgers. Edu/WSSP/StudentScholars/project/introduction/worms.html. Last accessed 12/05/2014 Wormatlas. (ND). Excretory System. Available: http://www.wormatlas.org/hermaphrodite/excretory/Excframeset.html. Last accessed 12/05/2014. Read More