Currently, the Aquatic Vertebrate Platform, supported by Junta de Andalucía, aims to facilitate investigation in aquatic vertebrate models to both national and international researchers and companies. To this end, the scientific manager of the facility, Ana Fernández-Miñán, offers advice on genomic experimentation with the aquatic vertebrates: zebrafish, medaka and Xenopus.

Specifically, services (please click next to download the list of AVP_SERVICES.pdf), collaborations and advice are offered on the following techniques *:

A.  General Transgenesis

To build expression vectors à la carte we use:

 A.1. The Tol2kit based on the multisite Gateway technology (1).

Fig: Tol2kit

 A.2. pTransgenesis vectors: compatible with Xenopus, zebrafish and medaka (2).

Fig: pTransgenesis vectors

In addition to keep a detailed record of the generated zebrafish transgenic lines, the Platform will also be in charge of their maintenance and distribution to interested researchers. Transgenic lines are available at CABD's Enhancer Screen.

B.  Embryology and Imaging

For the experimental testing of your transgenic lines, the following techniques are available:

 B.1.    Gain and loss-of-function studies by mRNA and morpholinos microinjection.

Morpholinos design and order information can be found at GENE TOOLS.

 B.2.    Tailored protocols for RNA in situ hybridization and inmunohistochemistry.

Fig: RNA in situ hybridization and inmunohistochemistry tailored protocols Fig: RNA in situ hybridization and inmunohistochemistry tailored protocols

 B.3.    in vivo confocal microscopy (time lapse).

In collaboration with the Microscopy Facility at CABD (see rates here).



 B.4.    Mosaic analysis by transplantation and mosaic injection.

Fig: Mosaic analysis

C.  Genomics

We also offer a range of options for genomic analysis:

 C.1. Enhancer activity assessment using the Zebrafish Enhancer Detector (ZED) vector (4).

Fig: ZED vector

The ZED vector consists of:
a) GFP under the control of a minimal promoter;
b) Insulator sequences to shield from positional effects;
c) Positive control for transgenesis (Cardiac Actin RFP).

 C.2. Analysis of the insulating capacity of genetic elements.

An efficient vector designed to test insulator’s functionality in zebrafish F0 injected embryos is used (4).

Fig: Analysis of the insulating capacity of genetic elements

Insulator’s activity is shown by the ability of the element to downregulate the strong Z48 brain enhancer from the cardiac actin promoter.

 C.3. Chromosome Conformation Capture (3C) (5) and 4c-seq technologies (6-8).

Fig: 3C-qPCR

Use of the 3C-qPCR technique to confirm distant interaction between two genomic positions, i.e. an enhancer and a promoter

4C-seq technology combines 3C with next generation sequencing. It allows to map the physical interactions' landscape of a given site of interest in a genome-wide manner.

Fig: 4C

Fig: Chip-PCR and Chip-seq

 C.4. Chip-PCR and Chip-seq techniques (9, 10).

Chromatin inmunoprecipitation using specific antibodies + massive sequencing

C.5. Basic bioinformatic support on Chip-seq experiments.

MACS (v.1.3.3) algorithm (Peak calling tool)

Gene Ontology Browser

The Aquatic Vertebrate Platform also offers a Guest Laboratory where to develop collaboration projects under the technical advice from the scientific manager of the facility.

* A, B1 and B2 services are free of charge for Consolider members. Rates for the rest of the services or rates for non-Consolider members must be discussed individually.

Should you be interested in any of the Platform services or in a collaboration, please contact the scientific manager.