Fly Transgenesis

The CONGENTO fly transgenesis service for the production of genetically modified flies relies on two main facilities (Champalimaud and Instituto Gulbenkian de Ciência) that work in a collaborative structure to improve timing and efficiency in the execution of your requests and is coordinated by Leonardo Guilgur (also available as a consultant if discussion of any project is required).

Supported by highly specialized HR – including postdoctoral scientists – and state-of-the-art equipment, we are prepared not only to supply the microinjection service but also to perform all the steps to generate customized construct cloning, if required, meaning that we offer an all-inclusive service to deliver researchers gene-edited flies. Besides regular p-element and Phi31 insertions, we are able to perform different editing requests, such as gene disruption, reporter knock-in, Tag knock-in, and seamless editing.

You can contact the service through fly@congento.org .

Services

P-element or PhiC31 transgenics

Basic

for customers that would like to do their own screening, we offer a Drosophila embryo injection service for p-element integrations into the genome randomly (classic p-element insertions) or with landing site (PhiC31 integrase-mediated transgenesis). Accurately purified DNA sample of the construct for injection is provided either by the customer or is purified by our team as a service option. Prior to injection, we check DNA concentration and mix with the helper plasmid (provided by us). Basic service includes injection of the construct into 200 embryos and sending back the survival larvae in vials. You are responsible for identifying your own transformants and balancing them.

Extended

we will rear the injected larvae to adulthood following the Basic services. Cross the G0 injected flies to virgins or males. Identify and collect the transformants. The collected transformant flies (at least 3 independent lines) will be sent to the customer and original vials will be maintained in our lab for about two weeks. You are responsible for balancing them.

Complete

in addition to obtaining transgenics for your constructs, we balance as many lines as you would require. We also offer mapping service for the case of a p-element random insertion (by inverse PCR).


dsRNAi generation lines

The approach used (see also TRiP lines here) is to generate transgenic animals with an RNAi hairpin under UAS-Gal4 control. The hairpin-containing transgenes are inserted via site-specific recombination into genomic loci known to be optimal for expression.

Basic

customer provides the dsRNA probes cloned, and we inject in the landing site required to produce the TRIP line. Basic service includes injection of the construct into 200 embryos and sending back the survival larvae in vials. You are responsible for identifying your own transformants and balancing them.

Extended

customer provides the dsRNA probes cloned, and we inject in the landing site required to produce the TRIP line. The collected transformant flies (at least 3 independent lines) will be sent to the customer and original vials will be maintained in our lab for about two weeks. You are responsible for balancing them.

Complete

in addition to obtaining transgenics RNAi line, we also offer balance as many lines as you would require.


CRISPR/Cas9 edited flies

With CRISPR technologies, we are able to modify an endogenous gene locus by insertions, deletions, or mutations as well as, by supplying a donor repair template, precisely edit genomic sequence or insert exogenous DNA (e.g. a specific TAG).

Basic (injection + screening)

for the injection and screening service the donor sequences and gRNAs must be provided cloned by the customer. We then perform the injection service selecting the most suitable protocol for your particular case: -DNA plasmid (gRNA) + DNA plasmid (Cas9) (with or without donor). -DNA plasmid (gRNA) + Cas9 protein purified (with or without donor). -DNA plasmid (gRNA) (with or without donor) into Drosophila stains that expresses Cas9 endogenously (this is the most efficient way nowadays). After the injection procedure we will confirm the genome-edited fly strains by sequencing that genome region.

Complete

this service includes designing and cloning guide RNA targets and recombination arms into donor vectors, DNA preparation, injection and screening to bring you gene-edited flies. We discuss with you the best strategy and then we do all the steps involved in generating the CRISPR modified flies by using Cas9 expressing lines. The editing type includes target gene disruption, reporter knock-in and tag knock-in.


#Discuss with us about your particular objective, we are able to do all the cloning procedures and also phenotypic characterization (e.g. in a particular tissue or developmental stage).



Other

DNA Cleaning service

We can clean your DNA using our in-house technique. This step will dramatically improve the survival rate and transformation efficiency.

DNA Preparation

You can send us DNA, LB plate, LB-Stab stab, or Glycerol stock for this service. If you send DNA, transformation has an extra cost.

Customized Cloning Service

Design and construct cassette for your specific research needs.

PCR Screening Service

Individual F1 progeny is back-crossed to balancer. We then perform single-fly genomic PCR on each fertile F1 flies.

Digestion Screening

Individual F1 progeny is back-crossed to balancer. We then perform single-fly genomic PCR on each fertile F1 flies. PCR products will be purified and cut by endonuclease and analyzed by gel electrophoresis.

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