Identification, analysis, and linkage mapping of expressed sequence tags from the Australian sheep blowfly

Abstract

Background: The Australian sheep blowfly Lucilia cuprina (Wiedemann) (Diptera: Calliphoridae) is a destructive pest of the sheep, a model organism for insecticide resistance research, and a valuable tool for medical and forensic professionals. However, genomic information on L. cuprina is still sparse.

Results: We report here the construction of an embryonic and 2 larval cDNA libraries for L. cuprina. A total of 29,816 expressed sequence tags (ESTs) were obtained and assembled into 7,464 unique clusters. The sequence collection captures a great diversity of genes, including those related to insecticide resistance (e.g., 12 cytochrome P450s, 2 glutathione S transferases, and 6 esterases). Compared to Drosophila melanogaster, codon preference is different in 13 of the 18 amino acids encoded by redundant codons, reflecting the lower overall GC content in L. cuprina. In addition, we demonstrated that the ESTs could be converted into informative gene markers by capitalizing on the known gene structures in the model organism D. melanogaster. We successfully assigned 41 genes to their respective chromosomes in L. cuprina. The relative locations of these loci revealed high but incomplete chromosomal synteny between L. cuprina and D. melanogaster.

Conclusions: Our results represent the first major transcriptomic undertaking in L. cuprina. These new genetic resources could be useful for the blowfly and insect research community.

Figure 1

An overview of the acquisition, assembly, analysis, and application of L. cuprina-expressed sequence tags. A total of 29,816 ESTs from embryonic and larval libraries was assembled into 7,464 unique sequence clusters using the TGICL procedures. E-values from BLAST searches were arranged in ascending order from left to right, indicated by the darkness of shade.

Figure 2

Size distribution of the 7,464 non-redundant sequence clusters.

Figure 3

Frequency distribution of contig sizes.

Figure 4

Mapping anchor loci by scoring intron length polymorphisms. Top right: Mapping pedigrees were generated using a backcross design initiated using the laboratory strain M_I5 and the field strain Tara. Top left: Primers were first tested in the 4 backcross parents to identify intron length polymorphism. Bottom: Informative primer pairs were used to screen the backcross pedigree (TMM01).

Figure 5

Chromosomal synteny between L. cuprina and D. Melanogaster. Genomic locations of the anchor loci are shown on the 6 D. melanogaster chromosome arms (X, 2L, 2R, 3L, 3R, and 4). The corresponding chromosomal origins (II, III, IV, V, and VI) of the Lucilia homologs are indicated in brackets. Six instances of synteny violation are shown in red.