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Describe the 16SrDNA analysis and how it is interpreted with respected bacterial phylogentic - Essay Example

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Running head: Description of the 16SrDNA analysis and how it is interpreted with respected bacterial phylogenic Institution of Affiliation: Date: The 16SrDNA analysis It was shown in laboratories of Woese amongst others that phylogenetic relationship pertaining and, indeed, the entire forms of life could be determined via comparing a stable portion of the genetic code…
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Describe the 16SrDNA analysis and how it is interpreted with respected bacterial phylogentic
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"Describe the 16SrDNA analysis and how it is interpreted with respected bacterial phylogentic"

The 16SrRNA gene has the ability to be compared not just among the entire bacteria but also can be compared with archeobacteria’s 16SrRNA and eucayotes’ 18SrRNA gene. With this understanding we can now delve into 16SrRNA gene sequence analysis. A commercial system (such as ABI; PrepMan DNA extraction reagent) or a standard method is used in extracting bacteria genomic DNA from whole cells. For the PCR the DNA is applies as a template to amplify a segment ranging about 500 or 1,500 bp relating to 16SrRNA gene sequence. Wide based or conventional primers complementary to preserved areas are utilized so that the area can be amplified from a given bacteria. The products of the PCR are purified to get rid of excess nucleotides and primers; a number of good commercial kits are present (e.g., Microcon-100 i.e. Microconncentrator columns [Millipore] and QiaQick PCR kit of purification [Qiagen]) (Jill, 2004). The following step is known as cycle sequencing. It is identical to PCR as it uses DNA (The first PCR cycle’s purified products) as the template. The two, reserve and forward sequences apply as templates in different reactions in which just the reverse or forward primer is used. A difference exist between PCR and Cycle sequencing as no new template is formed in the latter and the product involves a mixture of various lengths of DNA. This is attained by adding particularly labeled bases known as dye terminator, which, when incorporated randomly in this second cycle, they terminate the sequence (Janda & Abbott, 2007). Therefore, fragments of each magnitude are generated. Because every one of the four extra labeled terminator bases has dissimilar fluorescent dye, every of them absorbs at a dissimilar wavelength, the terminal base relating to every fragment can be determined via a fluorometer. Products are purified to get rid of unincorporated dye terminators, and using capillary electrophoresis or gel electrophoresis we determine the length of each product. Because we then understand the length as well as terminal base relating to every fragment, the bases’ sequences can be determined. The DNA’s two strands are sequenced dissimilarly, generating both reverse and forward (complementary) sequences (Janda & Abbott, 2007). The electropherogram, used for tracing detection of the fragments separated as they elute out of the column in which every base is represented via a dissimilar color, can be automatically or manually edited. It is probable to have fragments of several lengths thus well separated which each base of a 500-bp series can be determined. Electropherogram’s visual reediting can be used to resolve most of the ambiguities when they occur. How the 16SrDNA analysis is interpreted with respected bacterial phylogenic It is important that the quality of sequences is examined prior to using them. For instance, in a study that of strains of Actinomyces, sequences from Public database (GenBank-EMBL), a commercial database and the researcher’s internally generated databases were compared (Lang, Darling, & Eisen, 2013). The kind Actinomyces turicensis strain in the GenBank database as well as the 12 clinical strains which were sequenced indicated minimal (not any to two) dissimilarities as well as no ambiguous bases, showing that all were sequences of good quality. The sequence of the kind A. Read More
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