Microsatellite evolution: Mutations, sequence variation, and homoplasy in the hypervariable avian microsatellite locus HrU10

Abstract

BackgroundMicrosatellites are frequently used genetic markers in a wide range of applications, primarily due to their high length polymorphism levels that can easily be genotyped by fragment length analysis. However, the mode of microsatellite evolution is yet not fully understood, and the role of interrupting motifs for the stability of microsatellites remains to be explored in more detail. Here we present a sequence analysis of mutation events and a description of the structure of repeated regions in the hypervariable, pentanucleotide microsatellite locus HrU10 in barn swallows (Hirundo rustica) and tree swallows (Tachycineta bicolor).ResultsIn a large-scale parentage analysis in barn swallows and tree swallows, broods were screened for mutations at the HrU10 locus. In 41 cases in the barn swallows and 15 cases in the tree swallows, mutations corresponding to the loss or gain of one or two repeat units were detected. The parent and mutant offspring alleles were sequenced for 33 of these instances (26 in barn swallows and 7 in tree swallows). Replication slippage was considered the most likely mutational process. We tested the hypothesis that HrU10, a microsatellite with a wide allele size range, has an increased probability of introductions of interruptive motifs (IMs) with increasing length of the repeated region. Indeed, the number and length of the IMs was strongly positively correlated with the total length of the microsatellite. However, there was no significant correlation with the length of the longest stretch of perfectly repeated units, indicating a threshold level for the maximum length of perfectly repeated pentanucleotide motifs in stable HrU10 alleles. The combination of sequence and pedigree data revealed that 15 barn swallow mutations (58%) produced alleles that were size homoplasic to other alleles in the data set.ConclusionOur results give further insights into the mode of microsatellite evolution, and support the assumption of increased slippage rate with increased microsatellite length and a stabilizing effect of interrupting motifs for microsatellite regions consisting of perfect repeats. In addition, the observed extent of size homoplasy may impose a general caution against using hypervariable microsatellites in genetic diversity measures when alleles are identified by fragment length analysis only.

What phylogeny and gene genealogy analyses reveal about homoplasy in citrus microsatellite alleles

“…In principle, homoplasy is tightly linked to the mechanisms that cause mutations that produce new alleles, and hence, homoplasy is also coupled to the underlying mutation model (Lia et al 2007). The amount of homoplasy in microsatellite alleles is generally thought to increase with increasing time of divergence among taxa (van Oppen et al 2000) and increasing allele size since longer repeats are less stable than shorter repeats (Anmarkrud et al 2008). Previous interspecific studies of the sequence content of microsatellite alleles have revealed prevalent homoplasy including loss of the targeted repeat motif; hence, it has been suggested that these markers may not be useful for phylogenetic analysis above the species level (Ochieng et al 2007;Tesfaye et al 2007).…”

“…However, because homoplasy is thought to increase with time of divergence, it is also possible that the divergence time between species and genera has not been extensive enough to allow mutations to accumulate, and thus, significantly alter the sequence of these microsatellite alleles. Another possible explanation could be that these microsatellites are somewhat stable in distant citrus taxa due to their small size since larger repeat motifs have been shown to have more hidden sequence motifs than shorter alleles (Anmarkrud et al 2008). Consequently, one may be able to control or reduce the amount of homoplasy in microsatellite alleles by intentionally selecting microsatellites with shorter repeat elements.…”

“…On the other hand, the SMM assumes that there is equal probability of gaining or losing a single repeat unit within the microsatellite region to produce new distinguishable alleles. Unlike the IAM, SMM takes into account mutations back to a previous state (Anmarkrud et al 2008). The SMM assumes that mutations result in alleles that have similar repeat units to the alleles from which they were derived.…”

“…Two molecular evolution models commonly used for the analysis of SSR markers are the stepwise mutation model (SMM) and the infinite allele model (IAM). The IAM postulates that each new mutation produces a unique allele (Kimura and Crow 1964), which allows for the creation of an infinite number of allelic states not already present in the population (Estoup and Cornuet 1999;Anmarkrud et al 2008). On the other hand, the SMM assumes that there is equal probability of gaining or losing a single repeat unit within the microsatellite region to produce new distinguishable alleles.…”

Assessment of microsatellites in estimating inter- and intraspecific variation among Neotropical Crocodylus species

“…Because they are hypervariable and occur ubiquitously, microsatellites have widely been used as tools for a variety of fields of study such as population and conservation genetics, molecular ecology (mating behavior and gene flow), and wildlife DNA forensic analyses (Glenn et al, 1998;Fitzsimmons et al, 2001;Davis et al, 2001Davis et al, , 2002Dever et al, 2002;Avise, 2004;Anmarkrud et al, 2008).…”