Exploring Size Selection Methods to Optimize Read Length in 454 Sequencing

Mentor: Danielle McCarthy

Next generation DNA sequencing technologies such as 454 have replaced the traditional method of Sanger sequencing, but result in shorter lengths of the sequence reads generated during sequencing. Lisa explored variations on the process of preparing DNA for 454 sequencing, in order to find conditions that would produce optimal read lengths.

Lisa focused on optimizing the sizes of the DNA fragments sent to sequencing, and she therefore chose to test variations of the DNA shearing and size selection steps of DNA preparation. For 4 conditions, the samples Lisa prepared went through the entire 454 sequencing process, and for 18 other conditions, Lisa analyzed the samples to determine yield and size of the DNA fragments. Lisa discovered conditions that provide the best yields, and other conditions that provide better size selectivity, when compared to the current standard 454 protocol. The conditions producing the best size selectivity and yield should produce the longest average read lengths, a theory to be tested in future experiments.

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Lisa, a senior at Brockton High School, researched conditions for achieving optimal DNA yield, size selection of DNA, and read length during 454 sequencing.