Full text loading...
Chapter 16 : How Many Genes Does a Cell Need?
Category: Genomics and Bioinformatics; Environmental Microbiology
Ebook: Choose a downloadable PDF or ePub file. Chapter is a downloadable PDF file. File must be downloaded within 48 hours of purchase
This chapter reviews, discusses, and proposes studies aimed at determining how many Mycoplasma genitalium genes are really necessary when cells are grown under ideal laboratory conditions. A list of the 100 M. genitalium protein-coding genes disrupted in the 2006 study is provided in this chapter. Using The Institute for Genomic Research (TIGR) functional classification scheme, the 100 genes can be broken into 15 groups based on their main roles. Seven DNA repair and recombination genes were disrupted, including recA, which is one of the most ubiquitous proteins found in nature. A minimal set of genes cannot be convincingly determined by either comparative genomics or global transposon mutagenesis. However, these studies do identify more than 200 genes for core functions such as DNA replication, transcription, protein translation, energy metabolism, transport, lipid metabolism, nucleotide metabolism, and protein fate, which one has confidence will be represented in the minimal cell. There are three possible approaches to making a minimal cell: (i) cumulative inactivation of genes using mutagens that produce frameshifts, (ii) sequential genome reduction using recombineering methods, and (iii) chemical synthesis of a minimal genome and installation into receptive cell cytoplasm. A minimal genome may not by itself be useful in a practical sense, but one can envision in the not too distance future the ability to design and synthesize microbes for useful purposes such as production of pharmaceutical products, industrial compounds, and fuels. In the future, stripped down organisms outfitted with useful biosynthetic pathways could provide the basis for new industrial processes.
Comparison of orthologous genes in M. genitalium and H. influenzae Rd ( Mushigian and Koonin, 1996 ). The total number of protein-coding genes in each bacterium is based on the original 1995 annotations, which have been modified several times in the intervening years.
Evolutionary tree showing the relatedness of 13 mycoplasmas. P. asteris is an obligate intracellular organism and is thus distinct from others.
Diagram illustrating transposon mutagenesis. The transposon carries a transposase enzyme, an antibiotic resistance gene (TetR), and terminal sequences recognized by the transposase during transposition. The transposon is generally carried on a plasmid that does not replicate in the cell to be mutagenized. Antibiotic resistance is acquired by the cells only when the transposon jumps into the cell chromosome. A gene is disrupted when the transposon lands inside a gene.
M. genitalium genome map showing the location of genes and transposon insertion sites.
Deletion of a gene by recombineering using the lambda red system. Gene A is flanked by 50 bp of sequence on each side that are the targets for a linear piece of DNA that consists of an expressible galK gene flanked by the two 50-bp sequences. After introduction of the galK DNA fragment by electroporation, recombination occurs within the two 50-bp homologous segments, thus deleting gene A and replacing it with galK. Subsequent steps, as described in the text, counterselect for removal of the galK gene, leaving a clean (“scarless”) deletion. The process can be repeated many times to remove other genes.
Assembly of multiple overlapping pieces of DNA by in vitro recombination. If vector DNA is included in the reaction and it overlaps sequences at each end of the assembly, then a circular DNA is formed, which can be cloned in a suitable host cell.
A diagram illustrating the basic tenet of synthetic biology. In analogy to computers, the genome of a cell is the operating system and the cytoplasm of the cell is the hardware that runs the operating system.
Minimal protein-coding gene set defined by comparing M. genitalium and H. influenzae Rd ( Mushegian and Koonin, 1996 )
Summary of global transposon mutagenesis insertion site data
The essential and nonessential genes of M. genitalium are listed by main role and subroles using the TIGR functional classification system a