Possible ecological risks of transgenic organism release when transgenes affect mating success: Sexual selection and the Trojan gene hypothesis

Widespread interest in producing transgenic organisms is balanced by concern over ecological hazards, such as species extinction if such organisms were to be released into nature. An ecological risk associated with the introduction of a transgenic organism is that the transgene, though rare, can spread in a natural population. An increase in transgene frequency is often assumed to be unlikely because transgenic organisms typically have some viability disadvantage. Reduced viability is assumed to be common because transgenic individuals are best viewed as macromutants that lack any history of selection that could reduce negative fitness effects. However, these arguments ignore the potential advantageous effects of transgenes on some aspect of fitness such as mating success. Here, we examine the risk to a natural population after release of a few transgenic individuals when the transgene trait simultaneously increases transgenic male mating success and lowers the viability of transgenic offspring. We obtained relevant life history data by using the small cyprinodont fish, Japanese medaka (Oryzias latipes) as a model. Our deterministic equations predict that a transgene introduced into a natural population by a small number of transgenic fish will spread as a result of enhanced mating advantage, but the reduced viability of offspring will cause eventual local extinction of both populations. Such risks should be evaluated with each new transgenic animal before release.

Abundance of copepods from multinet samples during POLARSTERN cruise ANT-XX/1

Abundance and species composition of copepods were studied during the expedition ANT XXI/1 on a latitudinal transect in the eastern Atlantic from 34°49.5' N to 27°28.1' S between 2-20 November 2002. Stratified zooplankton tows were carried out at 19 stations with a multiple opening-closing net between 300 m water depth and the surface. Cyclopoid and calanoid copepods showed similar patterns of distribution and abundance. Oithona was the most abundant cyclopoid genus, followed by Oncaea. A total of 149 calanoid copepod species were identified. Clausocalanus was by far the most abundant genus, comprising on average about 45% of all calanoids, followed by Calocalanus (13%), Delibus (9%), Paracalanus (6%), and Pleuromamma (5%). All other genera comprised on average less than 5% each, with 40 genera less than 1%. The calanoid copepod communities were distinguished broadly in accordance with sea surface temperature, separating the subtropical from the tropical stations, and were largely determined by variation in species composition and species abundance. Nine Clausocalanus species were identified. The most numerous Clausocalanus species was C. furcatus, which on average comprised half of all adult of this genus. C. pergens, C. paululus, and C. jobei, contributed an average of 19%, 9%, and 9%, respectively. The Clausocalanus species differed markedly in their horizontal and vertical distributions: C. furcatus, C. jobei, and C. mastigophorus had widespread distributions and inhabited the upper water layers. Major differences between the species were found in abundance. C. paululus and C. arcuicornis were biantitropical and were absent or occurred in very low numbers in the equatorial zone. C. parapergens was found at all stations and showed a bimodal distribution pattern with maxima in the subtropics. C. pergens occurred in higher numbers only at the southern stations, where it replaced C. furcatus in dominance. In contrast to the widespread species, the bulk of the C. paululus, C. arcuicornis, C. parapergens, and C. pergens populations was concentrated in the colder, deeper water layers below the thermocline, thereby avoiding the warm surface waters. C. lividus was found only at the most northern and C. ingens only at the most southern stations. Both species were found almost exclusively in the upper 50 m. The distinct differences in abundance and horizontal and vertical distribution suggest a strong ecological differentiation among the Clausocalanus species.

Zebrafish KLF4 is essential for anterior mesendoderm/pre-polster differentiation and hatching

Gene knockout studies of Kruppel-like factors (KLFs) in mice have shown essential roles in organogenesis. A screen for KLF family members in zebrafish identified many KLFs. One of these, zebrafish KLF4 (zKLF4) is the homologue of neptune, a Xenopus laevis KLF. zKLF4 is expressed from approximately 80% epiboly a patch of dorsal/anterior mesendodermal cells called the pre-polster and, subsequently, in the polster and hatching gland. Here we investigate the function of zKLF4 using morpholino-based antisense oligonucleotides. Knockdown of zKLF4 resulted in complete absence of hatching gland formation and subsequent hatching in zebrafish. In addition, there was early knockdown of expression of the pre-polster/anterior mesendoderm markers CatL, cap1, and BMP4. These results indicate zKLF4 is expressed within the pre-polster, an early mesendodermal site, and that it plays a critical role in the differentiation of these cells into hatching gland cells.

Expression of Bf/C2 Gene(S) in the Japanese Medaka Fish

Complement factor B and C2 are two central serine proteases of the alternative and classical complement pathways, respectively, that serve as the catalytic subunits of the C3 convertase. Research has been completed using a female Japanese medaka fish, (Oryzias latipes), and other teleost and elasmobrach species in order to isolate eDNA clones and perform linkage analysis of the Bf/C2 gene(s). To further analyze the evolution of the complement system in teleosts, different tissues than the ones from previous studies of medaka fish were analyzed for the constitutive gene expression of factor B and C2. Bf/C2 sequences were amplified by reverse transcription-polymerase chain reaction with primers corresponding to the common amino acid sequences shared by mammalian Bf and C2. Agarose gel electrophoresis was used to visualize sample bands and to calculate the concentration of gene expression of the Bf/C2 gene(s) in each tissue. All five tissue types, kidney, liver, muscle, testis, and spleen from a male medaka fish demonstrated Bf/C2 gene(s) expression, confirming that the messages of Bf/C2 gene(s) are distributed throughout the medaka fish. Tissues of the spleen, liver, and kidney contained the highest concentrations of expression of Bf/C2 gene( s ), while tissues of the muscle and testis contained the lowest concentrations. This research also determined that RT-PCR allowed for more sensitive analysis of gene expression than other molecular biology techniques such as Northern blotting analysis.

PCR-Based Analysis of Mitochondrial DNA Copy Number, Mitochondrial DNA Damage, and Nuclear DNA Damage.

Because of the role that DNA damage and depletion play in human disease, it is important to develop and improve tools to assess these endpoints. This unit describes PCR-based methods to measure nuclear and mitochondrial DNA damage and copy number. Long amplicon quantitative polymerase chain reaction (LA-QPCR) is used to detect DNA damage by measuring the number of polymerase-inhibiting lesions present based on the amount of PCR amplification; real-time PCR (RT-PCR) is used to calculate genome content. In this unit, we provide step-by-step instructions to perform these assays in Homo sapiens, Mus musculus, Rattus norvegicus, Caenorhabditis elegans, Drosophila melanogaster, Danio rerio, Oryzias latipes, Fundulus grandis, and Fundulus heteroclitus, and discuss the advantages and disadvantages of these assays.