DNA hybridization studies

DNA/DNA hybridization studies indicate a taxonomic revision for these three species is in order. Even though these species produce spores (unpublished results) they bear no genetic similarity to C. thermocellum. 

In this section, we review basic procedures that are used in single-copy DNA hybridization studies. In addition to the PB/hydroxyapatite and SI/TEACL systems, we also discuss other methods such as the PERT system and the PT system that combines TEACL with hydroxyapatite. 

Since microarrays are simply spots of DNA arrayed in a known pattern, they also can be used for DNA-DNA hybridization studies. This is the basis for using micro-arrays for comparative DNA hybridizations, single nucleotide polymorphism (SNP) discovery, and chromatin immunoprecipiation followed by microarray chip hybridization (ChIP on chip analyses) to identify transcription factor binding sites (discussed below). 

A crucial aspect of virtually all DNA-DNA hybridization studies is that they involve only so-called single-copy DNA (scDNA) or "unique DNA. Total genomic DNA is made single-stranded by heating and then allowed to reassociate into duplex DNA after cooling. Repeated copies of DNA sequences re-anneal faster than single-copy sequences due to their higher concentration in the solution. Thus, under controlled conditions and time, it is possible to remove excess copies of repeated sequences. 

Various parameters derived from DNA-DNA hybridization studies can be used as measures of the genetic divergence between the taxa being compared. The most commonly used is the change in the median melting temperature, Tj. This is ususally given as AT which is the Tj of the homoduplex minus the T of the heteroduplex. While this type of measure is very useful, especially with complex eukaryotic genomes, matters can become more complex. 

DNA-DNA hybridization studies of the carnivorous marsupials I. The intergeneic relationships of bandicoots (Marsupialia Perameloldea). J Mol Evol (in press)... 

Table 3. DNA-DNA Hybridization Studies with Parthenogenetic Drosophila...

HiUiard, L., Zhao, X., and Tan, W., Immobilization of oligonucleotides onto silica nanoparticles for DNA hybridization studies. Anal. Chim. Acta, 2002,470,51-56. 

Brucker-Lea, C. J., M. S. Stottlemyre, D. A. Holman et al. 2000. Rotating rod renewable microcolumns for automated, solid-phase DNA hybridization studies. Anal. Chem. 72 4233-4141. 

In the following, a few examples for the use of surface plasmon fluorescence spectroscopy in DNA hybridization studies will be given. The general architecture of the sensor surface layer has already been introduced and is summarized in Figure 11. 

Immobilization of oligonucleotides onto silica nanopartides for DNA hybridization studies. Analytica Chimica Acta, 470, 51-6. 

The DNA of any given AAV serotype will hybridize approximately 33% with the DNA of anj of the other serotypes as judged by DNA RNA hybridization assays using nitrocellulose filters (Rose et al, 1968). Further studies by Koczot (1970) using competitive hybridization have shown that the related sequences are the same for all four serotypes. More recent competitive DNA DNA hybridization studies have indicated that the homology among the different serotypes is closer to 50% (Koczot, personal communication). 

Yeast mitochondrial DNA occurs as double-stranded 26-/im closed circles, a molecular size corresponding to about 50 x 10 daltons. The number of circles per mitochondrion may range from zero to about five the total amount of cellular mtDNA per wild-type cell varies with the strain and accounts for 10-25% of the total cellular DNA. RNA-DNA hybridization studies indicate that yeast mtDNA contains one cistron of each of the 15 S and 21 S RNA species and probably 20 tRNA cis-trons. It has been reported that mitochondria from HeLa cells contain only 12 tRNA cistrons, 9 on the heavy DNA strand and 3 on the light strand. These authors suggested that since the proteins formed by mitochondrial ribosomes are enriched in hydrophobic amino acids, an array of 12 tRNAs may be sufficient for the complete synthesis of the inner-membrane proteins by mitochondria. Alternatively, some nuclear coded tRNAs may be available to the mitochondrial protein-synthesizing system. 

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