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Modified Organism
2nd Generation Friendly™ Aedes aegypti
Record information and status
Record ID
115346
Status
Published
Date of creation
2020-01-08 16:20 UTC (austein.mcloughlin@cbd.int)
Date of publication
2020-01-08 16:20 UTC (austein.mcloughlin@cbd.int)

Living Modified Organism identity
The image below identifies the LMO through its unique identifier, trade name and a link to this page of the BCH. Click on it to download a larger image on your computer. For help on how to use it go to the LMO quick-links page.

LMO name
2nd Generation Friendly™ Aedes aegypti
Transformation event
OX5034
Developer(s)
Oxitec Limited
71 Milton Park
OX14 4RX
Oxford, England
Phone:+44 (0) 1235 832393
Fax:+44 (0) 1235 861138
Email:info@oxitec.com
Url:http://www.oxitec.com
Description
Aedes aegypti mosquitoes were modified female-specific lethality for population suppression and red fluorescence for visual detection.

Sex-specific lethality is achieved through the use of alternative splicing and the expression of a synthetic tetracycline‐transcriptional activator (tTAV) variant. In females, splicing occurs such that the tTAV coding sequence is translated. In males, alternative splicing causes several stop codons to be included upstream of the tTAV coding sequence, interrupting translation and preventing expression of tTAV. In the presence of tetracycline (rearing conditions), tTAV preferentially binds tetracycline instead of the tetracycline operator, thus transcription is repressed and occurs at a basal level. In the absence of tetracycline (environmental conditions), tTAV binds the operator sequences to promote high levels of transcription. High levels of tTAV expression is toxic as it prevents the cells from producing other transcripts required for normal functioning and results in lethality. Thus, females can be produced in rearing conditions, but fail to develop in environments without the presence of tetracycline. Due to the alternative splicing mechanism, males carry and transmit the self-limiting genes to progeny without requiring tetracycline. Population suppression is thus achieved through the biased selection of male offspring.

The modified mosquitoes also include the Discosoma sp. DsRed2 protein for red fluorescence under yellow light for visual detection of modified organisms. In comparison to OX513A mosquitoes (see "Related LMOs"), a synthetic nuclear localization signal was included to localize DsRed2 to the nuclei of cells, improving visualization (fluorescence) in larvae and adult mosquitoes.
Recipient Organism or Parental Organisms
The term Recipient organism refers to an organism (either already modified or non-modified) that was subjected to genetic modification, whereas Parental organisms refers to those that were involved in cross breeding or cell fusion.
Aedes aegypti - Yellow fever mosquito, AEDAE
Related LMOs
Dominant lethal Aedes aegypti mosquito
Dr. Lee Han Lim Conditional lethality Selectable marker genes and reporter genes
Show detection method(s)
Characteristics of the transformation process
Vector
pOX5034
Techniques used for the modification
  • Microinjection
Genetic elements construct
 
hr5 Transcriptional Enhancer
0.56 Kb
 
 
Immediate-early-1 gene promoter
0.63 Kb
 
 
Scraps intron
0.09 Kb
 
 
Nuclear localization signal
0.02 Kb
 
 
DsRed2 Fluorescent Protein
0.68 Kb
 
 
Nuclear localization signal
0.02 Kb
 
 
SV40 poly-adenylation signal
0.23 Kb
 
 
SV40 poly-adenylation signal
0.23 Kb
 
 
Tetracycline-controlled transactivator
1.01 Kb
 
 
Ubiquitin
0.23 Kb
 
 
Sex-specific splicing module
4.04 Kb
 
 
HSP70 minimal promoter
0.13 Kb
 
 
Tetracycline Operator
0.30 Kb
 
Further details
Notes regarding the genetic elements introduced or modified in this LMO
Fluorescent marker
Transcription of the fluorescent marker commences from the Autographa californica multiple nucleopolyhedrovirus immediate-early-1 gene promoter and terminates at the Simian vacuolating virus 40 (SV40) poly-adenylation signal. The transcript initially contains a Drosophila melanogaster scraps intron, two synthetic nuclear localization signals (nls) and the Discosoma sp. DsRed2 fluorescent protein. The scraps intron improves mRNA stability and is required for translation of the protein, but is not expected to be included in the translated polypeptide. After splicing and translation, the nls allows DsRed2 to transit and accumulate in the nuclei of cells.

Sex-specific lethality
Transcription of the Escherichia coli tetracycline-controlled transactivator (tTAV) commences from the D. melanogaster heat shock protein 70 minimal promoter and terminates at the SV40 poly-adenylation signal. The transcript contains the Aedes aegypti sex-specific intron, D. melanogaster ubiquitin and tTAV.  In females, splicing removes the sex-specific intron. The remaining transcript is translated to produce a poly-peptide containing ubiquitin and tTAV. Post-translation, ubiquitin is removed, leaving the functional tTAV. Ubiquitin is expected to enhance protein expression within insect cells. In males, splicing occurs from a further downstream site within the sex-specific splicing module. The resulting transcript then contains several stop codons upstream of ubiquitin and tTAV, which disrupt translation and thus prevents the production of the tTAV protein.

Transcriptional expression is also influenced by the tetracycline operator, which is adjacent to the tTAV cassette and acts as a repressible switch. In the presence of tetracycline, tTAV preferentially binds tetracycline rather than the operator sequences. Thus, transcription remains at a basal level and repressed. In the absence of tetracycline, tTAV binds the operator sequences and stimulates transcription, resulting in elevated levels of the tTAV protein.
LMO characteristics
Modified traits
  • Selectable marker genes and reporter genes
  • Female-specific lethality
Common use(s)
  • Biocontrol
Detection method(s)
Additional information
Modified mosquitoes can be detected by red fluorescence under yellow light (583 nm) due to the presence of DsRed2 protein. Fluorescence is expected to be localized to the nuclei of cells.
Additional Information
Additional Information
Male mosquitoes contain insecticide-susceptibility genes, which are expected to dilute insecticide resistance in wild populations.

In Portuguese, the modified mosquito is known as "Aedes do Bem™".
Other relevant website address or attached documents