Illumina Sequencing Technology - Report Example

Illumina Sequencing Technology Introduction The IIlumina sequencing technology is the most widely adopted and successful next-genertion sequencing (NGS) technology in the world (Lander, Linton, & Nusbaum, 20011). Reagents of Illumina sequencing support massively parallel sequencing with the use of a propriety method. A propriety method helps detect single bases as they get incorporated into the DNA growing strands (Lander, Linton, & Nusbaum, 20011). This technology is often applied in bacterial communities within the mangrove wetlands to generate the overlapped V6 tages from the end reads of Illumina pair. This paper sought to discuss the Illumina technology. It focused on how the technology works, how it was applied to solve a problem in bacteria community in mangrove wetlands, and its strengths and weaknesses over culture-independent methods (16SrRNA gene clone libraries, denaturing and temperature fragment length polymorphism (DGGE), and terminal restriction fragment length polymorphism). How the technology works In principle, the key concept behind the SBS technology is similar to the Capillary electrophoresis (CE) based Sanger sequencing. Fragment of the DNA identified from a signal that is emitted when each small fragrant gets re-synthesized from a template of DNA strand. SBS extends this principle cross multiple reactions in a massive parallel fashion. It is not limited to a few or single DNA fragments. This advancement makes it possible for a fast sequencing of stretches of base pairs of the DNA to span the entire genomes. The latest instruments have the capability of producing millions of data in one sequencing run (NIH News Release NHGRI, 2004). How it was applied to solve a particular problem? In this study, the researcher obtained 1,294,216 raw 16SrRNAV6 sequences from 12 sediment samples that had been collected from 4 different locations in the same mangrove land. The barcoded Illumina paired-end sequencing method was then used in generating the overlapped V6 tages from the end reads of Illumina pair. The pairedend reads were then overlapped with a view to obtain high-quality sequences of V6 tag. Overlapped sequences with ambiguous bases, sequences with errors in either reverse or forward primer, sequences that had variable tags longer than 90 bp and shorter than 50 bp an d sequences with two or more mismatches within the region of 46 and 48 bp during the overlap step were discarded. After barcoded Illumina paired-end sequencing, 779, 569 full V6 tags length remained. Chimera sequences were then removed using UCHIME. A total of 777,123 clean tags were obtained. Each sample had an average of 64,760.25 plus or minus 9,017.20 tags per sample. What are the strengths AND weaknesses of the new technology relative to one other? When used in bacterial communities within the mangrove wetlands, Illumina method has advantageous but also disadvantageous to other methods. As has been revealed in this study, studies on microbial ecology and mangrove microbial communities often follow three historical period. In these three periods different methods are applicable. These are the culture-dependent method, the culture-independent methods (16SrRNA gene clone libraries, denaturing and temperature fragment length polymorphism (DGGE), and terminal restriction fragment length polymorphism), and pyrosequencing and Illumina methods. Like, pyrosequencing, Illumina methods has an advantage over the other earlier methods in that though the first methods were successful in isolating large number of microbial strain, this early method of studying the microbial ecology became weakened by the phenomenon referred to as the “great plate count”. When compared to the next generation methods including 16SrRNA gene clone libraries, denaturing and temperature fragment length polymorphism (DGGE), and terminal restriction fragment length polymorphism, Illumina was advantageous in that unlike the aforementioned methods which could not successfully reveal details of highly diverse bacterial communities among the mangrove wetlands, Illumina method, could reveal a more detailed picture of microbial communities among the mangrove wetlands. Other advantages of using this technology include its wide application preparation methods. As opposed to other aforementioned technologies, Illumina technology is widely applicable in de novo sequencing, RNA sequencing for applications such as small RNA analysis and trascriptome, protein-nucleic acid interaction analysis, whole-genome sequencing, DNA sequencing, and candidate region target sequencing. It also supports both paired-and end and read libraries (Leamon, Lee, Lanza, & J et al, 2003). The third advantage is that SBS technology is unique as it is the only technology that offers a high resolution genome sequencing. Fourthly, it offers a long-insert paired-end reads with the use of a similar robust chemistry to enable efficient sequence assembly, large-scale structural variations detection, and de novo detection. Last yet important, the combination of longer reads and shorter reads helps increase its ability to fully characterize any type of genome (NIH News Release, 2006). The major shortcoming of that limits the use of this technology especially in the de now sequencing is its associated short sequencing read lengths. There is need for a new technology that can combine this technology with long read lengths in order to achieve electrophoresis based \sanger-sequences. This would enable quick and high production of quality de now genome sequencing (NIH News Release, 2006). Conclusion: The Ilumina sequencing technology is the most widely adopted and successful next-genertion sequencing (NGS) technology in bacterial communities within the mangrove wetlands in the world. This is owed to its attributed characteristics that make it advantageous to other techniques. Firstly, unlike, the other earlier methods (16SrRNA gene clone libraries, denaturing and temperature fragment length polymorphism (DGGE), and terminal restriction fragment length polymorphism), which could not successfully reveal details of highly diverse bacterial communities among the mangrove wetlands, Illumina technology can reveal a more detailed picture of microbial communities among the mangrove wetlands. Secondly, unlike other technologies, these technology has wide application. Illumina technology is widely applicable in de novo sequencing, RNA sequencing for applications such as small RNA analysis and trascriptome, protein-nucleic acid interaction analysis, whole-genome sequencing, DNA sequencing, and candidate region target sequencing. It also supports both paired-and end and read libraries. Thirdly SBS technology is unique as it is the only technology that offers a high resolution genome sequencing. Fourthly, it offers a long-insert paired-end reads with the use of a similar robust chemistry to enable efficient sequence assembly, large-scale structural variations detection, and de novo detection. Last yet important, the combination of longer reads and shorter reads helps increase its ability to fully characterize any type of genome References Lander, E., Linton, M., & Nusbaum, C. (2001). Nature 2001, 409, 860-921. Technologies. http:llwww.geneme.gov. Leamon, J. Lee, W., Lanza, J et al. (2003). Electrophoresis, 24, 3769-3777. NIH News Release NHGRI (October 2004). Seeks Next Generetion of Sequencing NIH News Release. (August 2006). NHGRI Expands Effort to Revolutionalize Sequencing Technologies. http://www.genome.gov Read More