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Development and evaluation of male-only strains of the Australian sheep blowfly, Lucilia cuprina.

Scott MJ - BMC Genet. (2014)

Bottom Line: From the 1960s to the 1980s there was a major effort to develop "field female killing" or FFK strains of L. cuprina that could be used for a cost-effective genetic control program.Males did not die in the field as normal copies of the eye color genes had been translocated to the Y chromosome and an autosome.Although the FFK strains showed some promise in field tests, a genetic control program in mainland Australia was never implemented for several reasons including instability of the FFK strains during mass rearing.

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ABSTRACT
The Australian sheep blowfly Lucilia cuprina (Wiedemann) is a major pest of sheep in Australia and New Zealand. From the 1960s to the 1980s there was a major effort to develop "field female killing" or FFK strains of L. cuprina that could be used for a cost-effective genetic control program. The FFK strains carried eye color mutations that were lethal to females in the field but not under conditions in the mass rearing facility. Males did not die in the field as normal copies of the eye color genes had been translocated to the Y chromosome and an autosome. Although the FFK strains showed some promise in field tests, a genetic control program in mainland Australia was never implemented for several reasons including instability of the FFK strains during mass rearing. A stable transgenic strain of L. cuprina that carried one or more dominant repressible female lethal genes offered the potential for efficient genetic control of blowfly populations. Here I review our research on tetracycline-repressible female lethal genetic systems, Lucilia germ-line transformation and sex determination genes that ultimately led to the successful development of transgenic "male-only" strains of L. cuprina. The technology developed for L. cuprina should be directly transferable to other blowfly livestock pests including L. sericata and the New World and Old World screwworm. 29.

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Comparison of the genomic organization and sex-specific transcripts of the L. cuprina and C. hominivorax transformer genes. The diagrams represents the Lctra (A) and Chtra genes (B). The exons are shown as square boxes, with exons in red representing common exons to both female and male mRNAs. Exons m1 and m2 in blue represent male specific exons. Introns are represented by solid lines. Exon and intron sizes are indicated. Red vertical lines represent the position of putative TRA/TRA2 binding sites within the male exon and the first intron. The male and female transcripts are shown below the genes. The 5' and 3' untranslated regions are represented by black boxes. Vertical black lines mark translational start and stop sites and in-frame translational stop sites in the male exons are marked with asterisks. The predicted lengths of the proteins encoded by the transcripts are indicated. Lctra and Chtra have a very similar exon-intron arrangement and sex-specific splicing pattern. Modified from [37].
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Figure 3: Comparison of the genomic organization and sex-specific transcripts of the L. cuprina and C. hominivorax transformer genes. The diagrams represents the Lctra (A) and Chtra genes (B). The exons are shown as square boxes, with exons in red representing common exons to both female and male mRNAs. Exons m1 and m2 in blue represent male specific exons. Introns are represented by solid lines. Exon and intron sizes are indicated. Red vertical lines represent the position of putative TRA/TRA2 binding sites within the male exon and the first intron. The male and female transcripts are shown below the genes. The 5' and 3' untranslated regions are represented by black boxes. Vertical black lines mark translational start and stop sites and in-frame translational stop sites in the male exons are marked with asterisks. The predicted lengths of the proteins encoded by the transcripts are indicated. Lctra and Chtra have a very similar exon-intron arrangement and sex-specific splicing pattern. Modified from [37].

Mentions: An alternative approach was to return to using introns from sex-specifically spliced transcripts to achieve female-specific gene expression at an earlier stage of development. Giuseppe Saccone and colleagues had shown that in the Mediterranean fruit fly, tra transcripts are sex-specifically spliced and splicing is autoregulated [34]. Moreover, RNAi-mediated knockdown of tra expression led to the transformation of XX individuals to males. This suggested that it could be possible to make a repressible female-male transformation system, which modeling suggests could be a very effective means for population control [35]. However, isolation of the L. cuprina tra gene proved to be a formidable challenge as the tra gene was poorly conserved between Drosophila and medfly. A fragment of the L. cuprina tra gene was isolated using PCR with degenerate primer pairs based on conserved amino acid motifs [36]. Subsequent analysis found that only the female tra transcript codes for TRA protein. The major male transcript includes an additional exon with several in-frame stop codons (Figure 3). As in C. capitata, the presence of multiple predicted TRA/TRA2 binding sites within the sex-specifically spliced first intron strongly suggested that splicing was autoregulated. The tra gene was shown to be essential for female development as injection of tra double-stranded RNA into the posterior end of embryos led to the development of XX adults with male genitalia. More recently we have isolated the tra gene from L. sericata, C. hominivorax and C. macellaria [37]. The overall organization of the tra genes from the four blow fly species is remarkably conserved, with a similar exon-intron arrangement and relative location of TRA/TRA2 binding sites (Figure 3). Lastly, we also isolated the L. cuprina dsx gene and showed that dsx transcripts are sex-specifically spliced as in Drosophila and housefly [38]. The presence of eight TRA/TRA2 sites in the female exon strongly suggested that dsx splicing in female is regulated by TRA as in Drosophila.


Development and evaluation of male-only strains of the Australian sheep blowfly, Lucilia cuprina.

Scott MJ - BMC Genet. (2014)

Comparison of the genomic organization and sex-specific transcripts of the L. cuprina and C. hominivorax transformer genes. The diagrams represents the Lctra (A) and Chtra genes (B). The exons are shown as square boxes, with exons in red representing common exons to both female and male mRNAs. Exons m1 and m2 in blue represent male specific exons. Introns are represented by solid lines. Exon and intron sizes are indicated. Red vertical lines represent the position of putative TRA/TRA2 binding sites within the male exon and the first intron. The male and female transcripts are shown below the genes. The 5' and 3' untranslated regions are represented by black boxes. Vertical black lines mark translational start and stop sites and in-frame translational stop sites in the male exons are marked with asterisks. The predicted lengths of the proteins encoded by the transcripts are indicated. Lctra and Chtra have a very similar exon-intron arrangement and sex-specific splicing pattern. Modified from [37].
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4255793&req=5

Figure 3: Comparison of the genomic organization and sex-specific transcripts of the L. cuprina and C. hominivorax transformer genes. The diagrams represents the Lctra (A) and Chtra genes (B). The exons are shown as square boxes, with exons in red representing common exons to both female and male mRNAs. Exons m1 and m2 in blue represent male specific exons. Introns are represented by solid lines. Exon and intron sizes are indicated. Red vertical lines represent the position of putative TRA/TRA2 binding sites within the male exon and the first intron. The male and female transcripts are shown below the genes. The 5' and 3' untranslated regions are represented by black boxes. Vertical black lines mark translational start and stop sites and in-frame translational stop sites in the male exons are marked with asterisks. The predicted lengths of the proteins encoded by the transcripts are indicated. Lctra and Chtra have a very similar exon-intron arrangement and sex-specific splicing pattern. Modified from [37].
Mentions: An alternative approach was to return to using introns from sex-specifically spliced transcripts to achieve female-specific gene expression at an earlier stage of development. Giuseppe Saccone and colleagues had shown that in the Mediterranean fruit fly, tra transcripts are sex-specifically spliced and splicing is autoregulated [34]. Moreover, RNAi-mediated knockdown of tra expression led to the transformation of XX individuals to males. This suggested that it could be possible to make a repressible female-male transformation system, which modeling suggests could be a very effective means for population control [35]. However, isolation of the L. cuprina tra gene proved to be a formidable challenge as the tra gene was poorly conserved between Drosophila and medfly. A fragment of the L. cuprina tra gene was isolated using PCR with degenerate primer pairs based on conserved amino acid motifs [36]. Subsequent analysis found that only the female tra transcript codes for TRA protein. The major male transcript includes an additional exon with several in-frame stop codons (Figure 3). As in C. capitata, the presence of multiple predicted TRA/TRA2 binding sites within the sex-specifically spliced first intron strongly suggested that splicing was autoregulated. The tra gene was shown to be essential for female development as injection of tra double-stranded RNA into the posterior end of embryos led to the development of XX adults with male genitalia. More recently we have isolated the tra gene from L. sericata, C. hominivorax and C. macellaria [37]. The overall organization of the tra genes from the four blow fly species is remarkably conserved, with a similar exon-intron arrangement and relative location of TRA/TRA2 binding sites (Figure 3). Lastly, we also isolated the L. cuprina dsx gene and showed that dsx transcripts are sex-specifically spliced as in Drosophila and housefly [38]. The presence of eight TRA/TRA2 sites in the female exon strongly suggested that dsx splicing in female is regulated by TRA as in Drosophila.

Bottom Line: From the 1960s to the 1980s there was a major effort to develop "field female killing" or FFK strains of L. cuprina that could be used for a cost-effective genetic control program.Males did not die in the field as normal copies of the eye color genes had been translocated to the Y chromosome and an autosome.Although the FFK strains showed some promise in field tests, a genetic control program in mainland Australia was never implemented for several reasons including instability of the FFK strains during mass rearing.

View Article: PubMed Central - HTML - PubMed

ABSTRACT
The Australian sheep blowfly Lucilia cuprina (Wiedemann) is a major pest of sheep in Australia and New Zealand. From the 1960s to the 1980s there was a major effort to develop "field female killing" or FFK strains of L. cuprina that could be used for a cost-effective genetic control program. The FFK strains carried eye color mutations that were lethal to females in the field but not under conditions in the mass rearing facility. Males did not die in the field as normal copies of the eye color genes had been translocated to the Y chromosome and an autosome. Although the FFK strains showed some promise in field tests, a genetic control program in mainland Australia was never implemented for several reasons including instability of the FFK strains during mass rearing. A stable transgenic strain of L. cuprina that carried one or more dominant repressible female lethal genes offered the potential for efficient genetic control of blowfly populations. Here I review our research on tetracycline-repressible female lethal genetic systems, Lucilia germ-line transformation and sex determination genes that ultimately led to the successful development of transgenic "male-only" strains of L. cuprina. The technology developed for L. cuprina should be directly transferable to other blowfly livestock pests including L. sericata and the New World and Old World screwworm. 29.

Show MeSH
Related in: MedlinePlus