Development of polymorphic microsatellite markers for Fagus pashanica (Fagaceae) using next-generation sequencing.

Fagus pashanica is an endangered and endemic tree species in China. To understand its genetic diversity and structure for effective conservation, we used next-generation sequencing data to develop a set of microsatellite markers. Twenty-three of the 68 designed loci were successfully amplified. Fifteen polymorphic loci with clear peaks were selected for further analyses in three F. pashanica populations sampled from Nanjiang, Wangcang and Pingwu counties in Sichuan Province, China. The number of alleles per locus ranged from two to 11. The levels of observed and expected heterozygosity ranged from 0.033-0.852 and 0.033-0.787, respectively. All 23 loci were also successfully amplified in F. longipetiolata and F. lucida, and 19 were successfully amplified in F. engleriana. These microsatellite markers will be useful for population genetic studies of F. pashanica and other Fagus species.

A comparison of reproductive isolation between two closely related oak species in zones of recent and ancient secondary contact.

BACKGROUND: Much of the debate over the evolutionary consequences of hybridization on genetic divergence and speciation results from the breakdown or reinforcement of reproductive barriers in secondary hybrid zones. Among hybrid populations established for different lengths of time following secondary contact, stronger reproductive barriers are generally expected to occur in zones with longer contact. However, in plants no detailed investigation of recent and ancient zones of secondary contact has been conducted despite the importance of such a comparative study. Here, we compare pre- and postzygotic reproductive barriers between two closely related oak species, Quercus mongolica and Q. liaotungensis, in such a situation. RESULTS: The recorded flowering times of both species overlapped in both contact zones. The fruit set at 10 and 30 days after interspecific hand pollination was not significantly lower than that after intraspecific pollination whenever Q. mongolica or Q. liaotungensis comprised the maternal parents in both populations. These results indicated that neither prezygotic phenological barriers nor interspecific incompatibility could have resulted in the reproductive isolation between the two species in both hybrid zones. However, the proportion of hybrid seeds produced by both species in the ancient zone was significantly lower than that recorded in the recent zone of secondary contact. In addition, the proportion of hybrid seeds simulated to form, assuming both random mating and an absence of postpollination barriers, was significantly higher than that detected in the ancient contact zone but not in the recent contact zone. These results suggest stronger early-acting postzygotic isolation between the two oak species in the ancient relative to the recent contact zone. CONCLUSIONS: Our comparative study demonstrated that postzygotic barriers during seed maturity were the main contributing factor to total reproductive isolation, particularly in the ancient contact zone, which aided species delimitation. In the recently formed secondary contact zone, pre- and postzygotic barriers were not well developed, and a high frequency of natural hybridization was evident. To our knowledge this study provides the first comparison of reproductive isolation between the ancient and recent secondary contact zones in plants and helps to clarify the evolutionary consequences of hybridization in a temporal context.

Temperature responses of mutation rate and mutational spectrum in an Escherichia coli strain and the correlation with metabolic rate.

BACKGROUND: Temperature is a major determinant of spontaneous mutation, but the precise mode, and the underlying mechanisms, of the temperature influences remain less clear. Here we used a mutation accumulation approach combined with whole-genome sequencing to investigate the temperature dependence of spontaneous mutation in an Escherichia coli strain. Experiments were performed under aerobic conditions at 25, 28 and 37 °C, three temperatures that were non-stressful for the bacterium but caused significantly different bacterial growth rates. RESULTS: Mutation rate did not differ between 25 and 28 °C, but was higher at 37 °C. Detailed analyses of the molecular spectrum of mutations were performed; and a particularly interesting finding is that higher temperature led to a bias of mutation to coding, relative to noncoding, DNA. Furthermore, the temperature response of mutation rate was extremely similar to that of metabolic rate, consistent with an idea that metabolic rate predicts mutation rate. CONCLUSIONS: Temperature affects mutation rate and the types of mutation supply, both being crucial for the opportunity of natural selection. Our results help understand how temperature drives evolutionary speed of organisms and thus the global patterns of biodiversity. This study also lend support to the metabolic theory of ecology for linking metabolic rate and molecular evolution rate.

Genome sequences of two closely related strains of Escherichia coli K-12 GM4792.

Escherichia coli lab strains K-12 GM4792 Lac(+) and GM4792 Lac(-) carry opposite lactose markers, which are useful for distinguishing evolved lines as they produce different colored colonies. The two closely related strains are chosen as ancestors for our ongoing studies of experimental evolution. Here, we describe the genome sequences, annotation, and features of GM4792 Lac(+) and GM4792 Lac(-). GM4792 Lac(+) has a 4,622,342-bp long chromosome with 4,061 protein-coding genes and 83 RNA genes. Similarly, the genome of GM4792 Lac(-) consists of a 4,621,656-bp chromosome containing 4,043 protein-coding genes and 74 RNA genes. Genome comparison analysis reveals that the differences between GM4792 Lac(+) and GM4792 Lac(-) are minimal and limited to only the targeted lac region. Moreover, a previous study on competitive experimentation indicates the two strains are identical or nearly identical in survivability except for lactose utilization in a nitrogen-limited environment. Therefore, at both a genetic and a phenotypic level, GM4792 Lac(+) and GM4792 Lac(-), with opposite neutral markers, are ideal systems for future experimental evolution studies.

Female reproductive success decreases with display size in monkshood, Aconitum kusnezoffii (Ranunculaceae).

BACKGROUND AND AIMS: Reduction in female fitness in large clones can occur as a result of increased geitonogamous self-fertilization and its influence through inbreeding depression. This possibility was investigated in the self-compatible, bee-pollinated perennial herb Aconitum kusnezoffii which varies in clone size. METHODS: Field investigations were conducted on pollinator behaviour, flowering phenology and variation in seed set. The effects of self-pollination following controlled self- and cross-pollination were also examined. Selfing rates of differently sized clones were assessed using allozyme markers. KEY RESULTS: High rates of geitonogamous pollination were associated with large display size. Female fitness at the ramet level decreased with clone size. Fruit and seed set under cross-pollination were significantly higher than those under self-pollination. The pre-dispersal inbreeding depression was estimated as 0.502 based on the difference in seed set per flower between self- and cross-pollinated flowers. Selfing rates of differently sized clones did not differ. CONCLUSIONS: It is concluded that in A. kusnezoffii the negative effects of self-pollination causing reduced female fertility with clone size arise primarily from a strong early-acting inbreeding depression leading to the abortion of selfed embryos prior to seed maturation.

Tetra: an improved program for population genetic analysis of allotetraploid microsatellite data.

Population genetic analysis of allotetraploid microsatellite data has lagged far behind that of diploid data, largely because of an inability to determine allele copy number for partial heterozygotes. tetrasat developed by Markwith et al. (2006) uses an iterative substitution process to account for all probable combinations of allele copy numbers in populations with partial heterozygote samples. However, tetrasat cannot deal with microsatellite data containing more than 15 partial heterozygotes, because of an exponential increase in genotype combinations. tetra can handle the microsatellite data containing infinite partial heterozygotes. In the program tetra, the frequencies of alleles are measured as the probability with which the known alleles occur in unknown allele locations. The Hardy-Weinberg expected heterozygosity and Nei's coefficient of gene differentiation are calculated based on allele frequencies. The mean and standard error of expected heterozygosity are estimated through bootstrap method.