Informative enzyme/probe combinations for the multilocus DNA fingerprinting of marsupials

Abstract

DNA fingerprinting has become an invaluable tool in the study of population genetics, paternity success, and individual identification; however, the species specificity of some methods has made the wide-range screening of many different species very time-consuming. In this study we describe the development and application of reliable and informative DNA fingerprinting techniques in a range of marsupial species using three different restriction enzyme and two oligonucleotide probe combinations. Six species from four marsupial families, the koala (Phascolarctidae), tammar wallaby (Macropodidae), southern hairy-nosed wombat (Vombatidae), kowari, and dusky and brown marsupial mice (Dasyuridae) were examined. Restriction enzymes HinfI AluI and HaeIII were used in combination with the digoxygenin (DIG)-labelled oligonucleotide probes (CAC)5 and (GGAT)4. The combinations of HinfI/(GGAT)4, AluI/ (CAC)5 and AluI/(GGAT)4 were the most informative, providing highly resolved bands, low background, and the lowest band sharing between individuals. The genetic diversity evident within the different species showed a clear relationship between the level of band sharing and population size. The greatest levels of band sharing were found in the kowaris (80%), which were part of a long-term captive colony originating from a few founders, and the lowest levels of band sharing were found in the marsupial mice (30-35%) and tammar wallaby (45%), which were caught from large outbred wild populations.

A study for minisatellitic markers of Conidiobolus coronatus’ pathogenicity to Galleria mellonella larvae

“…Our result of hybridization of fungal DNA with the human minisatellite 33.6 probe, together with the reports on the application of the human minisatellite probes to a wide variety of organisms, from plants to animals (Jeffreys et al, 1991;Lambert et al, 1997; Rogstad et al, 1988) and protozoa (Carnaby et al, 1995), demonstrates that some minisatellites are widespread among the eukaryotes and these sequences are conserved during evolution. Conidiobolus coronatus is an entomopathogenic fungus which kills host insects rapidly.…”

Sperm competition and genetic diversity in the agile antechinus (Dasyuridae: Antechinus agilis)

“…There has been much debate over the potential advantages to females of a multiply-sired litter and the increased levels of reproductive ®tness that result (Loman, Madsen & Hakansson, 1988;Arnqvist, 1989). Together these two outbreeding mechanisms could account for the observation of Lambert et al (1997) that, of seven marsupial species examined, A. agilis showed the highest level of genetic heterozygosity. Furthermore, the population advantages of multiplysired litters are perhaps of equal signi®cance for A. agilis in which all males die at the end of their ®rst breeding season.…”

“…Extracted DNA from each sample was ethanolprecipitated and quanti®ed by spectrophotometry (Lambert et al, 1997). Extracted DNA from each sample was ethanolprecipitated and quanti®ed by spectrophotometry (Lambert et al, 1997).…”

“…The DNA pro®ling techniques, speci®cally the restriction enzyme/probe combination, used in this study has been determined as the most informative (highest band variability) for marsupials (Lambert et al, 1997) and consequently enabled paternity assignment from unique male bands by inclusion (Fig. 2).…”

Sperm Transport and Storage in the Agile Antechinus (Antechinus agilis)1

“…This suggests that the order of copulation is an important factor in determining paternity success for male A. agilis. Efficient transport of spermatozoa to the oviduct and lengthy periods of storage are designed to ensure the success of multiple matings and encourage sperm competition and multiple-paternity litters, which may help to maintain the high levels of heterozygosity observed in wild A. agilis populations [37].…”