Repetitive genome elements in a European corn borer, Ostrinia nubilalis, bacterial artificial chromosome library were indicated by bacterial artificial chromosome end sequencing and development of sequence tag site markers: implications for lepidopteran genomic research

Abstract

The European corn borer, Ostrinia nubilalis, is a serious pest of food, fiber, and biofuel crops in Europe, North America, and Asia and a model system for insect olfaction and speciation. A bacterial artificial chromosome library constructed for O. nubilalis contains 36 864 clones with an estimated average insert size of >or=120 kb and genome coverage of 8.8-fold. Screening OnB1 clones comprising approximately 2.76 genome equivalents determined the physical position of 24 sequence tag site markers, including markers linked to ecologically important and Bacillus thuringiensis toxin resistance traits. OnB1 bacterial artificial chromosome end sequence reads (GenBank dbGSS accessions ET217010 to ET217273) showed homology to annotated genes or expressed sequence tags and identified repetitive genome elements, O. nubilalis miniature subterminal inverted repeat transposable elements (OnMITE01 and OnMITE02), and ezi-like long interspersed nuclear elements. Mobility of OnMITE01 was demonstrated by the presence or absence in O. nubilalis of introns at two different loci. A (GTCT)n tetranucleotide repeat at the 5' ends of OnMITE01 and OnMITE02 are evidence for transposon-mediated movement of lepidopteran microsatellite loci. The number of repetitive elements in lepidopteran genomes will affect genome assembly and marker development. Single-locus sequence tag site markers described here have downstream application for integration within linkage maps and comparative genomic studies. Résumé : La pyrale du maïs, Ostrinia nubialis, est un ravageur important des cultures pour des fins alimentaires, textiles et énergétiques en Europe, en Amérique du Nord et en Asie. De plus, il s'agit d'une espèce modèle pour l'olfaction et la spéciation chez les insectes. Une banque de chromosomes bactériens artificiels a été produite pour l'O. nubialis et totalise 36 864 dont la taille moyenne des inserts est d'environ 120 kb pour une couverture génomique de 8,8 fois. Un criblage de clones OnB1 conférant une couverture de 2,76 équivalents génomiques a permis de déterminer la position physique de 24 marqueurs de séquence connue dont des marqueurs liés à des caractères écologiques importants ainsi qu'à la résistance à la toxine du Bacillus thuringiensis. Les séquences des extrémités de clones chromosomes bactériens artificiels OnB1 (accessions GenBank dbGSS ET217010 à ET217273) présentaient de l'homologie avec des gènes annotés ou des étiquettes de séquences exprimées et ont permis d'identifier des séquences répétitives : des éléments transposables miniatures à répé-titions subterminales inversées (OnMITE01 et OnMITE02) ainsi que des éléments nucléaires dispersés de grande taille) de type ezi. La mobilité des éléments OnMITE01 a été démontrée par leur présence ou absence au sein d'introns chez deux locus de l'O. nubialis. Un motif tétranucléotidique répété (GTCT)n aux extrémités 5' des éléments OnMITE01 et On-MITE02 fournit la preuve de la mobilité de locus microsatellites chez les insectes via le mouvement de transposons. Le nombre d'élé...

A Helitron-Like Transposon Superfamily from Lepidoptera Disrupts (GAAA)n Microsatellites and is Responsible for Flanking Sequence Similarity within a Microsatellite Family

“…Isolation and Annotation of an Ostrinia nubilalis TE Portion of an O. nubilalis bacterial artificial chromosome (BAC) library (OnB1; 120 kb average insert size) was screened by PCR with oligonucleotide primers that amplified a 198 bp product specific for the mobile O. nubilalis MITElike element, OnMITE01 (Coates et al 2009). PCR products had estimated sizes ranging from 190-210 bp, and were observed from 63 of 96 (65.6%) OnB1 clones.…”

“…An O. nubilalis BAC library, OnB1, clones 04M1-04M24, 04B1-04B24, 55M1-55M24, and 50K1-50K24 were screened for OnMITE01 presence by PCR using primers OnMITE01-F 5 0 -TCC YAA CTA ATA TTA TAR ATG CGA AAG-3 0 and OnMITE01-R 5 0 -CCC GCG TGG AAT TTT GTC TG-3 0 (Coates et al 2009). PCR amplification of each BAC clone took place in 10 ll reaction volumes containing 1.5 mM MgCl 2 , 50 lM dNTPs, 5 ng BAC DNA, 1.8 pmol of each primer, 2 ll 59 thermal polymerase buffer (Promega), and 0.3125U GoTaq DNA polymerase (Promega, Madison, WI, USA).…”

“…Oligonucleotide primers OnMINE-1-F (5 0 -CAT TTA TTG CCA TGG ACA YCA C-3 0 ) and OnMINE-1-R (5 0 -GTG CGA CAG GGT GGC ACT-3 0 ) were designed from alignment of OnMINE-1 and OnB1 BAC end sequence (BESs) from GenBank GSS accessions ET217118 and ET217119 (Coates et al 2009). The 139 bp predicted PCR fragment was PCR amplified under conditions outlined for primers OnMITE01-F and -R, except the 72°C extension time was decreased to 10 s and products were separated by 2% agarose gel electrophoresis.…”

“…MITEs lack an internal protein coding region, and mobility is mediated by trans-acting factors encoded by related autonomous TEs (Dufresne et al 2006). MITEs are found ubiquitously within eukaryotic genomes, including mosquito (Tu 2000;Quesneville et al 2006), Drosophila (Locke et al 1999;Vivas et al 1999;Miller et al 2000;Wilder and Hollocher 2001;Yang and Barbash 2008), and Lepidoptera (Chen and Li 2007;Coates et al 2009) and have a role in altering genome structure including insertion at microsatellite loci (Akagi et al 2001).…”

“…Genome associations between TEs and microsatellite loci are also observed when integrations occur preferentially into tandem repeats (Akagi et al 2001;Temnykh et al 2001) or when tandem repeats hitchhike within mobile elements (Wilder and Hollocher 2001;López-Giráldez et al 2006;Coates et al 2009). Modification of microsatellite loci by TE insertion has been documented.…”