Identification of restriction-fragment-length polymorphisms in genomic DNA of the lesser snow goose (Anser caerulescens caerulescens)

A genomic library of partially EcoRI-digested DNA from the lesser snow goose, Anser caerulescens caerulescens, was constructed in the phage vector Charon 4. Phage containing only unique sequences were identified by screening plaques with 32P-labeled genomic DNA. Restriction-fragment- length polymorphisms (RFLPs) were identified by probing DNA from 11-13 male birds from the breeding colony at La Perouse Bay. Of the 17 probes examined, all detected RFLPs with at least one of EcoRi, HindIII, Msp1, and Taq1. Several of them identified highly variable regions with multiple alleles. These RFLPs are valuable DNA markers that can be used for (1) the examination of DNA variation, relatedness, and genetic distance and (2) assessing paternity and maternity. These data suggest that there are higher levels of variation of DNA sequence in birds than had previously been thought to exist.      

Captive breeding, paternity determination, and genetic variation in chimpanzees (Pan troglodytes) in the Australasian region

DNA “fingerprinting” using polymorphic (CA)-repeat microsatellite markers was used to quantify the level of genetic variation present in chimpanzees (Pan troglodytes) in the Australasian region. These markers were also used to determine the paternity of chimpanzees born at Taronga Zoo over a 20-year period. The results suggested that the dominant male in the colony was responsible for siring most, but not all, of the offspring. Where the dominant male was excluded from paternity, the sire was identifiable if all candidate males were available for typing. This enabled us to prove the captive origin of offspring born in the colony during this period. Thus, microsatellite analysis was a useful tool for assignment of familial relationships and improving genetic management of breeding colonies.     

DNA “fingerprinting” and the genetic management of a captive chimpanzee population (Pan troglodytes)

DNA fingerprinting probes are cloned sequences which simultaneously detect a large number of similar hypervariable loci in the target DNA. The resulting highly polymorphic pattern visualized on an autoradiograph allows resolution of questions concerning individual identification and parentage. M13 bacteriophage has been used as a DNA fingerprinting probe for paternity ascertainment among captive chimpanzees housed in multi-male groups as part of the National Chimpanzee Breeding and Research Program. In 31 cases of unknown paternity where DNA samples for mother, offspring, and all potential sires were available, DNA fingerprinting with M13 resulted in the unambiguous assignment of paternity for all 31 infants. Knowledge of pedigrees among the captive-born animals is used to address several issues important in the genetic management of captive breeding colonies, including estimation of effective population size and of the rate of decline in genetic variability, variance in male and female reproduction, and the effect of social dominance on male reproductive success. Our analysis demonstrates the beneficial effects of genetic management by comparing the managed dedicated cohort to the Bastrop colony as a whole.     

Genomic fingerprints of organisms from different taxonomic groups: the use of phage M13 DNA as a hybridization probe

Hypervariable polymorphic patterns were detected using wild-type M13 DNA as a probe in genomic DNAs of very different organisms ranging from procaryotes and lower eucaryotes to upper plants and animals, including human beings. Due to somatic stability of highly polymorphic patterns and their discrete inheritance, individual-specific restriction pattern analysis ("DNA fingerprinting") with this test probe was found to be useful in applied human genetics, in particular, for identifying paternity and maternity, and mapping of human genomes. The data obtained also demonstrate some possibilities of the DNA fingerprinting technology in genetics and selection of agricultural plants and animals, such as variety analysis, classification and registration of individual inbred lines and strains, as well as identification of bacterial strains.     

Patterns of multiple paternity and maternity in fishes

The characterization of patterns of multiple mating is a major facet of molecular ecology and is paramount to understanding the evolution of behaviours associated with parental care and mate choice. Over the last 15 years, fishes have been particularly well studied with respect to multiple maternity and paternity thanks to the widespread application of microsatellite markers. The present review focusses on the impressive literature on genetic parentage in fishes. In studies of natural populations, we find that multiple paternity is extremely common across fish species, whereas rates of multiple maternity are much more variable. In species with nest defence, for example, rates of multiple maternity are strongly bimodal, and the occurrence of multiple dams per brood is either rare or the rule. The sex of the care‐giving parent is correlated with the rate of multiple parentage: when males provide uniparental care, rates of multiple paternity are low compared to rates of multiple maternity; when females provide parental care, either alone or assisted by males, rates of multiple paternity are highly variable, whereas rates of multiple maternity are quite low. These patterns may reflect conflicts between the reproductive interests of males and females. We also find that fishes in which females brood the offspring internally display much higher rates of multiple paternity compared to mammals or birds, whereas reptiles are intermediate. Male‐nesting fish species, however, show rates of multiple paternity more similar to those found in other vertebrates. © 2011 The Linnean Society of London, Biological Journal of the Linnean Society, 2011, 103 , 735–760.     

First searchable database for DNA profiles of human cell lines: sequential use of fingerprint techniques for authentication.

The authenticity and freedom from cross-contaminants of a cell line are important prerequisites for any research, development or production programs involving cell lines. Mini- and microsatellites in the human genome harboring variable-numbers of tandem repeat (VNTR) DNA markers allow individualization at the DNA level and are of practical value for genetic linkage mapping, forensic legal medicine, paternity testing, monitoring of bone marrow transplants, and individualization of established cell lines. We have validated fingerprint techniques of different single- and multiple-locus VNTRs enabling the establishment of a searchable database of DNA profiles. As a result, multiplexed polymerase chain reaction amplification fragment length polymorphism (AmpFLP) of four prominent and highly polymorphic minisatellite VNTR loci was proven as the best tool for screening the uniqueness of DNA profiles in a fingerprint database. In order to avoid false positivity, identical or similar DNA profiles based on AmpFLP VNTR were tested further using a multi-locus fingerprint system. Our data demonstrate that misidentification remains a chronic problem among human continuous cell lines (detailed information at URL http://www.dsmz.de). The combination of rapidly generated DNA profiles based on single-locus VNTR loci, their authentication by screening the fingerprint database, and confirmation of duplicate banding patterns using multilocus fingerprints constitute a highly reliable and robust method, which enables high fidelity and quality of maintenance independent from the quantity of individual cell lines.     

DNA fingerprinting and the problems of paternity determination in an inbred captive population of guinea baboons (Papio hamadryas papio)

Multilocus DNA fingerprinting was carried out on 65 individuals from a captive colony of guinea baboons (Papio hamadryas papio) at Brookfield Zoo, in order to determine the allocation of reproductive success among 7 active males. DNA fingerprinting was found to reveal very low levels of genetic variability in the study population, rendering discrimination of different levels of relatedness, and hence paternity, impossible. A method was therefore developed for emphasizing the region of the fingerprint pattern which revealed the greatest level of band variability, and the effect of this experimental modification on band sharing statistics was tested. Band sharing coefficients among unrelated individuals were significantly lower using the modified system, which was then applied to paternity testing in the whole population. However even when using the modified system, of the 33 offspring analyzed only 4 could be assigned solely to 1 male, 14 offspring were assigned to 1 of 2 males, 7 offspring had 3 potential fathers, and the remainder had 4 or more possible fathers. The implications of the limitations of these data for behavioural studies and genetic management of captive populations are discussed.     

DNA分子标记在动物个体识别与亲权鉴定方面的应用

自从DNA分子标记技术建立以来,各种DNA分子标记相继被广泛地应用于遗传图谱的构建,评估遗传多样性以及个体识别,亲权鉴定等方面,综述了利用DNA分子标记技术,进行动物个体识别与亲权鉴定的原理,研究进展,现状及其应用前景与存在的问题。     

Evolutionary dynamics of microsatellite DNA

Within the past decade microsatellites have developed into one of the most popular genetic markers. Despite the widespread use of microsatellite analysis, an integral picture of the mutational dynamics of microsatellite DNA is just beginning to emerge. Here, I review both generally agreed and controversial results about the mutational dynamics of microsatellite DNA. Microsatellites are short DNA sequence stretches in which a motif of one to six bases is tandemly repeated. It has been known for some time that these sequences can differ in repeat number among individuals. With the advent of polymerase chain reaction (PCR) technology this property of microsatellite DNA was converted into a highly versatile genetic marker (Litt and Luty 1989; Tautz 1989; Weber and May 1989). Polymerase chain reaction products of different length can be amplified with primers flanking the variable microsatellite region. Due to the availability of high-throughput capillary sequencers or mass spectrography the sizing of alleles is no longer a bottleneck in microsatellite analysis. The almost random distribution of microsatellites and their high level of polymorphism greatly facilitated the construction of genetic maps (Dietrich et al. 1994; Dib et al. 1996) and enabled subsequent positional cloning of several genes. Almost at the same time, microsatellites were established as the marker of choice for the identification of individuals and paternity testing. The high sensitivity of PCR-based microsatellite analysis was not only of great benefit for forensics, but opened completely new research areas, such as the analysis of samples with limited DNA amounts (e.g., many social insects) or degraded DNA (e.g., feces, museum material) (Schl?tterer and Pemberton 1998). More recently, microsatellite analysis has also been employed in population genetics (Goldstein and Schl?tterer 1999). Compared with allozymes, microsatellites offer the advantage that, in principle, several thousand potentially polymorphic markers are available. Nevertheless, the application of microsatellites to population genetic questions requires a more detailed understanding of the mutation processes of microsatellite DNA as the evolutionary time frames covered in population genetics are often too long to allow novel microsatellite mutations to be ignored. Additional interest in the evolution of microsatellite DNA comes from the discovery that trinucleotide repeats, a special class of microsatellites, are involved in human neurodegenerative diseases (e.g., fragile X and Huntington's disease). A detailed understanding of the processes underlying microsatellite instability is therefore an important contribution toward a better understanding of these human neurodegenerative diseases.     

Extra-pair paternity in the shag, Phalacrocorax aristotelis as determined by DNA fingerprinting

The frequency of chicks resulting from extra-pair copulation in the shag, Phalacrocorax aristotelis , was measured by DNA fingerprinting. DNA fingerprints were taken from both sexes of 15 pairs and their chicks (28) in a subcolony on the Isle of May, UK. It was found that 18% of the chicks had extra-pair paternity, and one chick (3.5%) was not the offspring of either member of the pair, implying either a polygynous male whose second female was fertilized by another male or adoption. Although no observations of courtship and copulation were made in the same season, observations in a previous year on the same colony of shags showed that 14.1% of the copulations by males were not with the female with whom that male reared young. The similarities and differences are discussed between these results on the level of extra-pair copulations and of extra-pair paternity and those of other studies where both observations of extra-pair copulations and various measures of the degree of extra-pair paternity have been made.     

Sex-linked markers facilitate genetic parentage analyses in knobbed whelk broods

To explore the potential of sex-linked polymorphisms for genetic parentage analyses in natural populations, we have employed a recently discovered "X-linked" microsatellite marker (in conjunction with polymorphic autosomal loci) to deduce biological paternity and maternity for large numbers of encapsulated embryos within individual broods of the knobbed whelk (Busycon carica). Empirical findings illustrate how such sex-linked genetic tags can in special instances find at least three novel utilities in genetic dissections of large-clutch species: clarification of paternity assignments that had remained ambiguous from di-locus autosomal data alone; elucidation of linkage relationships among pairs of autosomal loci; and illumination of maternity (and thereby paternity also) in broods for which neither biological parent was known from independent evidence.     

Microsatellite DNA polymorphism confirms reproductive isolation and reveals differences in population genetic structure of cryptic pipistrelle bat species

Previous studies have indicated that the common European pipistrelle bat ( Pipistrellus pipistrellus ) comprises two cryptic species, P. pipistrellus and Pipistrellus pygmaeus , which differ in echolocation call frequency and mitochondrial DNA sequence. However, levels of divergence based on nuclear markers have not been examined, and hence the potential for male-mediated gene flow between the species cannot be discounted. Moreover, little is known about population structure and migration patterns in either species. Here, we describe the use of microsatellites to investigate nuclear DNA differentiation between, and the pattern of population genetic structure within, the two cryptic pipistrelle species. In total, 1300 individuals from 82 maternity colonies were sampled across the British Isles and Continental Europe. We show, using multivariate analyses, that colonies of the same species are generally genetically more similar to each other than to those from the other species regardless of geographical location. Our findings support the hypothesis that the species are reproductively isolated. Significant patterns of genetic isolation by distance were identified in both species, indicating that mating may occur before any long-distance autumnal migration. The presence of a sea channel does not confer higher levels of genetic differentiation among colonies over and above distance alone in either species. Differences in genetic population structure were identified between the species, with P. pipistrellus showing a wider range of levels of genetic differentiation among colonies and a stronger relationship between genetic and geographical distance than P. pygmaeus . Differences in dispersal, mating behaviour, colony size and/or postglacial colonization patterns could contribute to the differences observed.  © 2007 The Linnean Society of London, Biological Journal of the Linnean Society , 2007, 90 , 539–550.     

Genetic mating system and the significance of harem associations in the bat Saccopteryx bilineata

We analysed the polygynous mating system of the bat Saccopteryx bilineata using behaviour observations and genetic data on 11 microsatellite DNA loci. Basic social units in S. bilineata are harem groups that consist of single males and up to eight females. Colonies comprise several harem groups, and the composition of colonies and harems is often stable over several reproductive seasons. The combination of parentage exclusion and likelihood-based parentage assignment in this study produced detailed parentage information for a large colony of S. bilineata. Reproduction occurred mostly within the colony (17% extra-colony paternity), but social associations in harems within the colony did not represent reproductive units (70% extra-harem paternity). The latter finding was consistent over three reproductive seasons. Spatial association of the roosting sites of males and females could not explain parentage patterns in the colony. Even though intra-harem paternity was less frequent than expected, it contributed significantly to reproduction of harem males. On average, the number of offspring sired by a male with females in his harem territory increased significantly with harem size, which corresponds to the higher energetic investment that is related to the maintenance of large harems. However, extra-harem paternity was not correlated with a male's harem size or intra-harem reproductive success. This suggests that individual preferences of females rather than male traits associated with the ability to defend large harems are most likely to cause the detected differences between social association and genetic mating system.     

Calculation of probability of paternity using DNA sequences.

This paper formally incorporates allele measurement error into the Essen-Möller version of the probability of paternity. For highly polymorphic genetic systems, an approximate solution to the problem is developed resulting in simple formulas. The DNA sequence of the D14S1 region provided a practical example for testing this approximation. For these sequences, allelic uncertainty arises from determining length of DNA fragments from mobility in gel electrophoresis. D14S1 and standard test results from 35 paternity cases establish the validity of our computational method.     

Dissecting the DNA damage response using functional genomics approaches in S. cerevisiae

The faithful transmission of genetic information from a mother to daughter cells can only occur if the integrity of the genome is preserved. DNA transactions within cells are tightly regulated to prevent DNA damage. When events that challenge genome integrity do occur, a complex web of DNA damage response pathways comes into play. Studies in model organisms have contributed significantly to the understanding of these pathways. In the last decade, the development of functional genomics techniques in S.cerevisiae has ushered in systematic approaches for the study of complex cellular processes. These methods have enabled the systematic interrogation of the DNA damage response.     

Male reproductive success in a promiscuous mammal: behavioural estimates compared with genetic paternity

Molecular techniques have enabled behavioural ecologists to reassess mating systems from a genetic perspective. Studies of paternity frequently reveal that mating behaviour does not always reflect parentage, and may bring to light alternative mating tactics. Here we present a comparison of behavioural and genetic measures of male reproductive success in a mammalian mating system in which both sexes are highly promiscuous. Rather than having a stable harem social structure, Soay rams (Ovis aries) on the island of Hirta, St Kilda, UK usually consort with individual oestrous ewes sequentially. Not all matings occur between consort pairs, however, and ewes have been seen to mate with up to 10 different rams on the same day. Using locus-specific polymorphism at five protein and 10 microsatellite DNA loci, we determined paternity for 236 lambs born into three cohorts, and compared paternity with estimates of mating success derived from more than one census of rutting behaviour. The correlation between the number of ewes with which each ram was witnessed in consort and the number of paternities assigned was positive and statistically significant, and rams that were observed in consort with a ewe were 18 times more likely to have sired her offspring than other candidate rams. However, most lambs (73%) were not sired by a ram seen in consort with the oestrous mother. Many juveniles, yearlings and some adult rams were rarely seen in consort with ewes, yet were assigned a significant number of paternities. These results suggest that mating tactics differ between age groups, and that alternative mating strategies among adults that do not involve forming consorts with many females also confer mating success. For these reasons, census-based observations of consort associations between individuals cannot be used to accurately estimate individual male reproductive success in this population.     

Genetic analysis of a successful repatriation programme: giant Galápagos tortoises

As natural populations of endangered species dwindle to precarious levels, remaining members are sometimes brought into captivity, allowed to breed and their offspring returned to the natural habitat. One goal of such repatriation programmes is to retain as much of the genetic variation of the species as possible. A taxon of giant Galápagos tortoises on the island of Espa?ola has been the subject of a captive breeding-repatriation programme for 33 years. Core breeders, consisting of 12 females and three males, have produced more than 1200 offspring that have been released on Espa?ola where in situ reproduction has recently been observed. Using microsatellite DNA markers, we have determined the maternity and paternity of 132 repatriated offspring. Contributions of the breeders are highly skewed. This has led to a further loss of genetic variation that is detrimental to the long-term survival of the population. Modifications to the breeding programme could alleviate this problem.     

Colony genetic structure in the Australian jumper ant <Emphasis Type="Italic">Myrmecia pilosula</Emphasis>

Eusocial insects vary significantly in colony queen number and mating frequency, resulting in a wide range of social structures. Detailed studies of colony genetic structure are essential to elucidate how various factors affect the relatedness and the sociogenetic organization of colonies. In this study, we investigated the colony structure of the Australian jumper ant Myrmecia pilosula using polymorphic microsatellite markers. Nestmate queens within polygynous colonies, and queens and their mates, were generally unrelated. The number of queens per colony ranged from 1 to 4. Queens were estimated to mate with 1–9 inferred and 1.0–11.4 effective mates. This is the first time that the rare co-occurrence of polygyny and high polyandry has been found in the M. pilosula species group. Significant maternity and paternity skews were detected at the population level. We also found an isolation-by-distance pattern, and together with the occurrence of polygynous polydomy, this suggests the occurrence of dependent colony foundation in M. pilosula; however, independent colony foundation may co-occur since queens of this species have fully developed wings and can fly. There is no support for the predicted negative association between polygyny and polyandry in ants.     

Mitochondrial DNA (mtDNA) reveals that female Bechstein's bats live in closed societies

We present a microgeographic analysis of mitochondrial DNA (mtDNA) in Bechstein's bats using three sources of control region sequence variability, including a novel mtDNA microsatellite, to assess individual relatedness both within and among 10 maternity colonies. Comparison of marker variability among 268 adult females revealed little genetic variability within each colony. However, most colonies were clearly distinguished by colony-specific mitochondrial haplotypes (total n = 28). Low intracolony variability and strong haplotype segregation among colonies, was reflected by an extraordinary high FST of 0.68, indicating a very low intercolony dispersal rate of approximately one female in five generations. Haplotype distribution among 18 solitary males showed that males frequently disperse between colony locations, indicating the absence of dispersal barriers. Bechstein's bat maternity colonies are thus closed groups that comprise 20-40 females probably belonging to only one or, at most, two matrilines. The genetic population structure of Bechstein's bats is in agreement with the hypothesis that females seek familiar and, at least, partially related cooperation partners for raising their young. Alternatively strong philopatry might reflect the importance of profound roost or habitat knowledge for successful reproduction in female Bechstein's bats.