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metagenomics-based sequencing techniques. Metagenomic data obtained on sequencing of DNA,
mRNA or proteins isolated from natural samples is compared with the genomic or metabolic data
obtained from cultured strains that is available on the net (Fig. 3). Large scale DNA sequencing
strategies in which DNA extracted from cells/environmental samples is cloned into appropriate
vectors such as phagemids or bacterial artifi cial chromosome (BAC) and the libraries constructed
are used in sequencing (Fig. 4). These large fragments of DNA (up to 100 kb) contain several genes
arranged in a precise order in which they are found in the genomes they come from and in determining
the exact order of sequence of genes it is quite necessary to clone overlapping fragments (Fig. 5).
Alternatively, the extracted DNA either from the cells or environmental sequences is fragmented
into small pieces (approximately 3 kb) and is cloned into conventional vectors in a procedure called
“shotgun cloning”. These are subjcted to sequencing techniques. The sequences are then assembled
by using overlapping DNA regions. A number of new sequencing methods have been developed
that parallelly conduct the sequencing process leading to the production of thousands or millions
of sequences at once.
Figure 3:
Work fl ow for reconstructing metabolic models from metagenomic data. Courtesy U.S. Department of Energy
Genomic Science program and the website http://genomicscience.energy.gov.
Majority of the prokaryotic genomes have been sequenced at JCVI (http://www.jcvi.org,
founded by J. Craig Venter who spearheaded privately the sequencing of human genome project
that was accomplished in 2000 and discovered the shotgun sequencing method). JCVI coordinates
researches at four organizations, the Center for Advancement of Genomics, The Institute for Genome
Research, The Institute for Biological Energy Alternatives and the J. Craig Venter Science Foundation
Joint Technology Center and the JCVI has facilities in Rockville, Maryland and La Jolla, California.
The other sequencing Institutes are DOE-JGI (that unites the expertise of fi ve national laboratories:
Lawrence Berkeley, Lawrence Livemore, Los Alamos, Oak Ridge along with Hudson Alpha
Institute for Biotechnology and is operated by the University of California for the U.S. Department
of Energy), Gordon and Betty Moore Foundation Microbial Genome Sequencing Project (http://
www.moore.org), Kazusa DNA Research Institute, Kisarazu, Japan (http://www.kazusa.org) and
