Novel microsatellite markers for the oriental fruit mothGrapholita molesta(Lepidoptera: Tortricidae) and effects of null alleles on population genetics analyses

Abstract

The oriental fruit moth (OFM) Grapholita molesta (Lepidoptera: Tortricidae) is an important economic pest of stone and pome fruits worldwide. We sequenced the OFM genome using next-generation sequencing and characterized the microsatellite distribution. In total, 56,674 microsatellites were identified, with 11,584 loci suitable for primer design. Twenty-seven polymorphic microsatellites, including 24 loci with trinucleotide repeat and three with pentanucleotide repeat, were validated in 95 individuals from four natural populations. The allele numbers ranged from 4 to 40, with an average value of 13.7 per locus. A high frequency of null alleles was observed in most loci developed for the OFM. Three marker panels, all of the loci, nine loci with the lowest null allele frequencies, and nine loci with the highest null allele frequencies, were established for population genetics analyses. The null allele influenced estimations of genetic diversity parameters but not the OFM's genetic structure. Both a STRUCTURE analysis and a discriminant analysis of principal components, using the three marker panels, divided the four natural populations into three groups. However, more individuals were incorrectly assigned by the STRUCTURE analysis when the marker panel with the highest null allele frequency was used compared with the other two panels. Our study provides empirical research on the effects of null alleles on population genetics analyses. The microsatellites developed will be valuable markers for genetic studies of the OFM.

Multiple refugia from penultimate glaciations in East Asia demonstrated by phylogeography and ecological modelling of an insect pest

“…Alignment of the nucleotide sequences was checked and realigned following the guidance of amino acid alignment. For nuclear markers, 12 microsatellite loci developed from randomly sequenced genomic sequences were used [ 59 ] (Additional file 1 : Table S3). The microsatellites were genotyped using GENEMAPPER version 4.0 (Applied Biosystems).…”

Census and contemporary effective population size of two populations of the protected Spanish Moon Moth (Graellsia isabellae)

“…Despite having excluded locus GI18 from analyses involving Puebla, we cannot rule out a slight effect of null alleles, a well-known phenomenon in Lepidoptera (e.g. Song et al, 2017) in our results (see GI6 at Puebla/2008 and GI17 at Bujaruelo/ 2010, Table S3). Nevertheless, our conclusion of moderate-large N e for Puebla is not jeopardised by this possibility, as null alleles bias towards low N e when estimated with the LD method (Sved et al, 2013).…”

Genome-wide developed microsatellites reveal a weak population differentiation in the hoverflyEupeodes corollae(Diptera: Syrphidae) across China

“…79,138 microsatellite loci were isolated from the randomly sequenced genome sequences of E. corollae with 5000 (6.32%) dinucleotide repeat (DNR) sites, 29221 (36.92%) trinucleotide repeat (TNR) sites, 30988 (39.16%) tetranucleotide repeat (TTNR) sites, 6635 (8.38%) pentanucleotide repeats (PNR) sites and 7294 (9.22%) hexanucleotide repeat (HNR) sites. The frequency of dinucleotide repeats in E. corollae is unusually low when compared with other insect species such as Grapholita molesta [30] (Lepidoptera), Aphis glycines (Hemiptera) [38] and Obolodiplosis robiniae (Diptera) [39], which shows the distribution of microsatellites to vary among species [40, 41].…”

“…The recent development of genomic sequencing technology has made it relatively easy to isolate powerful microsatellites from large numbers of candidates at a genome-wide scale [42]. This method has been used in population structure analyses in many species, such as Grapholita molesta [30], Frankliniella occidentalis [43] and Carposina sasakii [29]. In our study, the 24 microsatellites developed are highly efficient in terms of amplification and polymorphism, enabling us to assess the population genetic structure of E. corollae .…”

“…QDD was used to isolate microsatellites and design primers [28]. The outputs of primer pairs from QDD were further filtered by the following criteria [29, 30] : (i) the corresponding microsatellites were pure and specific; (ii) the design strategy of ‘A’ was used to avoid primer secondary structure and repeats; (iii) the minimum distance between the 3′ end of a primer and its target region should be longer than 10 bp; (iv) the annealing temperature for each primer pairs was set between 58 °C and 62 °C to avoid large differences among primers; (v) the estimated PCR product size of the primer pairs was from 100 to 350 bp.…”