Dataset Information

Building early-larval sexing systems for genetic control of the Australian sheep blow fly Lucilia cuprina using two constitutive promoters.

ABSTRACT: Transgenic sexing strains (TSS) that carry conditional female lethal genes are advantageous for genetic control programs based on the sterile insect technique (SIT). It is desirable if females die early in development as larval diet is a major cost for mass production facilities. This can be achieved by using a gene promoter that is only active in embryos to drive expression of the tetracycline transactivator (tTA), the transcription factor commonly used in two-component TSS. While an embryo-specific promoter is ideal it may not be essential for assembling an effective TSS as tTA can be repressed by addition of tetracycline to the diet at larval and/or adult stages. Here we have investigated this idea by isolating and employing the promoters from the Lucilia spitting image and actin 5C genes to drive tTA expression in embryos and later stages. L. cuprina TSS with the tTA drivers and tTA-regulated tetO-Lshid effectors produced only females when raised on a limited tetracycline diet. The Lshid transgene contains a sex-specific intron and as a consequence only females produce LsHID protein. TSS females died at early larval stages, which makes the lines advantageous for an SIT program.

SUBMITTER: Yan Y

PROVIDER: S-EPMC5451413 | biostudies-literature | 2017 May

REPOSITORIES: biostudies-literature

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Building early-larval sexing systems for genetic control of the Australian sheep blow fly Lucilia cuprina using two constitutive promoters.

Yan Ying Y Linger Rebecca J RJ Scott Maxwell J MJ

Scientific reports 20170531 1

Transgenic sexing strains (TSS) that carry conditional female lethal genes are advantageous for genetic control programs based on the sterile insect technique (SIT). It is desirable if females die early in development as larval diet is a major cost for mass production facilities. This can be achieved by using a gene promoter that is only active in embryos to drive expression of the tetracycline transactivator (tTA), the transcription factor commonly used in two-component TSS. While an embryo-spe ...[more]

PMID: 28566730

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Project description:BackgroundThe sterile insect technique (SIT) has been successfully used in many pest management programs worldwide. Some SIT programs release both sexes due to the lack of genetic sexing strains or efficient sex separation methods but sterile females are ineffective control agents. Transgenic sexing strains (TSS) using the tetracycline-off control system have been developed in a variety of insect pests, from which females die by either of two commonly used lethal effectors: overexpression of the transcription factor tetracycline transactivator (tTA) or ectopic expression of a proapoptotic gene, such as head involution defective (hid). The lethality from tTA overexpression is thought to be due to "transcriptional squelching", while hid causes lethality by induction of apoptosis. This study aims to create and characterize a TSS of Lucilia cuprina, which is a major pest of sheep, by combining both lethal effectors in a single transgenic strain.ResultsHere a stable TSS of L. cuprina (DH6) that carries two lethal effectors was successfully generated, by crossing FL3#2 which carries a female-specific tTA overexpression cassette, with EF1#12 which carries a tTA-regulated LshidAla2 cassette. Females with one copy of the FL3#2 transgene are viable but up to 99.8% of homozygous females die at the pupal stage when raised on diet that lacks tetracycline. Additionally, the female lethality of FL3#2 was partially repressed by supplying tetracycline to the parental generation. With an additional LshidAla2 effector, the female lethality of DH6 is 100% dominant and cannot be repressed by maternal tetracycline. DH6 females die at the late-larval stage. Several fitness parameters important for mass rearing such as hatching rate, adult emergence and sex ratio were comparable to those of the wild type strain.ConclusionsCompared to the parental FL3#2 strain, the DH6 strain shows stronger female lethality and lethality occurs at an earlier stage of development. The combination of two tTA-dependent lethal effectors could improve strain stability under mass rearing and could reduce the risk of resistance in the field if fertile males are released. Our approach could be easily adapted for other pest species for an efficient, safe and sustainable genetic control program.

Dataset Information

Building early-larval sexing systems for genetic control of the Australian sheep blow fly Lucilia cuprina using two constitutive promoters.

Publications

Building early-larval sexing systems for genetic control of the Australian sheep blow fly Lucilia cuprina using two constitutive promoters.

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