|Genome Sequencing Section
DOE Human Genome Program Contractor-Grantee Workshop
9. Automation of Finishing at JGI-LLNL
Stephanie Stilwagen, Matt Nolan,
Andre Arellano, Karolyn Burkhart-Schultz, Long Do, Arthur Kobyashi, Brent
Kronmiller, Madison Macht, David Ow, Hoan Phan, Glenda Quan, Melissa Ramirez,
Warren Regala, Christina Sanders, Astrid Terry, Stephan Trong, Nelson Velasco,
Vijay Viswanathan, Anthony Carrano, and Jane Lamerdin
Lawrence Livermore National Laboratory has generated 10.5 Mb of highly accurate, finished genomic sequence of selected large insert clones (e.g. cosmid, BAC, P1) from chromosome 19 of which 8.6 Mb has been completed within the last fiscal year. We were able to achieve this eight-fold increase with the introduction of a suite of finishing tools and web-based computer interfaces which are directly linked to automated robotic workstations. The LLNL sequencing strategy utilizes a 'shotgun' approach to generate our initial sequence data. The next stage of the process is pre-finishing which involves the selection of clones for re-sequencing for initial gap closure and ambiguity resolution. We have automated pre-finishing by utilizing the LLNL developed software tool, Swedish to select clones for re-sequencing with either the dye primer or dye terminator chemistry. After one round of pre-finishing, a project moves to the finishing phase and is assigned to a finisher.
A finisher makes use of multiple software tools in an iterative manner to obtain contiguous sequence that meets our standards for double-strand coverage and sequence quality. Finishing involves closing the remaining gaps, resolving ambiguities, and validating the assembly. Automation of the finishing process makes use of Consed, Swedish, Finnish, web interfaces, and robotic workstations to increase efficiency and throughput. While these tools have had a significant impact on our productivity, additional tools and automation are still necessary to decrease the amount of human intervention required for finishing to meet the challenge of completing the Human Genome by 2003.
This work was performed by Lawrence Livermore National Laboratory under the auspices of the U.S. Department of Energy, Contract No. W-7405-Eng-48.
|Author Index||Sequencing Technologies||Microbial Genome Program|
|Search||Mapping||Ethical, Legal, & Social Issues|