viruses were injected to follicles on both wings for later studies. Chickens were raised in cages and observed on a daily basis over a two-month period. The regenerated feathers were plucked and examined with a dissection or scanning electron micrograph microscope for abnormalities compared with normal primary remiges. Histology and in situ hybridizationParaffin sections (5 mm) were stained with haematoxylin and eosin or prepared for in situ hybridization following routine procedures 26 . Cryostat sections (10 mm) were stained with X-gal. TUNEL staining was performed using a kit (Roche). Nonradioactive wholemount or section in situ hybridization or section in situ hybridization was performed according to the protocol described 22,26 . After hybridization, sections were incubated with an antidigoxigenin Fab conjugated to alkaline phosphatase (Boehringer Mannheim). Colour was detected by incubating with a Boehringer Mannheim purple substrate (Roche).

For the very short-period sdB eclipsing binary HW Vir, we present new CCD photometry made from 2000 through 2008. In order to obtain consistency of the binary parameters, our new light curves, showing sharp eclipses and a striking reflection effect, were analyzed simultaneously with previously published radial-velocity data. The secondary star parameters of M 2 =0.14 M ⊙ , R 2 =0.18 R ⊙ , and T 2 =3,084 K are consistent with those of an M6-7 main sequence star. A credibility issue regarding bolometric corrections is emphasized. More than 250 times of minimum light, including our 41 timings and spanning more than 24 yrs, were used for a period study. From a detailed analysis of the O-C diagram, it emerged that the orbital period of HW Vir has varied as a combination of a downward-opening parabola and two sinusoidal variations, with cycle lengths of P 3 =15.8 yr and P 4 =9.1 yr and semi-amplitudes of K 3 =77 s and K 4 =23 s, respectively. The continuous period decrease with a rate of −8.28×10 −9 d yr −1 may be produced by angular momentum loss due to magnetic stellar wind braking but not by gravitational radiation. Of the possible causes of the cyclical components of the period change, apsidal motion and magnetic period modulation can be ruled out. The most reasonable explanation of both cyclical variations is a pair of light-travel-time effects driven by the presence of two substellar companions with projected masses of M 3 sin i 3 =19.2 M Jup and M 4 sin i 4 =8.5 M Jup . The two objects are the first circumbinary planets known to have been formed in a protoplanetary disk as well the first ones discovered by using the eclipse-timing method. The detection implies that planets could be common around binary stars just as are planets around single stars and demonstrates that planetary systems formed in a circumbinary disk can survive over long time scales.Depending on the thermal inertia of their massive atmospheres, the hemispheres of the planets turned toward the stars can experience substantial reciprocating temperature changes during the minutes-long primary eclipse intervals.

Strong evidence exists for polyploidy having occurred during the evolution of the tribe Brassiceae. We show evidence for the dynamic and ongoing diploidization process by comparative analysis of the sequences of four paralogous Brassica rapa BAC clones and the homologous 124-kb segment of Arabidopsis thaliana chromosome 5. We estimated the times since divergence of the paralogous and homologous lineages. The three paralogous subgenomes of B. rapa triplicated 13 to 17 million years ago (MYA), very soon after the Arabidopsis and Brassica divergence occurred at 17 to 18 MYA. In addition, a pair of BACs represents a more recent segmental duplication, which occurred ;0.8 MYA, and provides an exception to the general expectation of three paralogous segments within the B. rapa genome. The Brassica genome segments show extensive interspersed gene loss relative to the inferred structure of the ancestral genome, whereas the Arabidopsis genome segment appears little changed. Representatives of all 32 genes in the Arabidopsis genome segment are represented in Brassica, but the hexaploid complement of 96 has been reduced to 54 in the three subgenomes, with compression of the genomic region lengths they occupy to between 52 and 110 kb. The gene content of the recently duplicated B. rapa genome segments is identical, but intergenic sequences differ.

A detailed genetic linkage map of Brassica rapa has been constructed containing 545 sequence-tagged loci covering 1287 cM, with an average mapping interval of 2.4 cM. The loci were identified using a combination of 520 RFLP and 25 PCR-based markers. RFLP probes were derived from 359 B. rapa EST clones and amplification products of 11 B. rapa and 26 Arabidopsis. Including 21 SSR markers provided anchors to previously published linkage maps for B. rapa and B. napus and is followed as the referenced mapping of R1-R10. The sequence-tagged markers allowed interpretation of the pattern of chromosome duplications within the B. rapa genome and comparison with Arabidopsis. A total of 62 EST markers showing a single RFLP band were mapped through 10 linkage groups, indicating that these can be valuable anchoring markers for chromosome-based genome sequencing of B. rapa. Other RFLP probes gave rise to 2-5 loci, inferring that B. rapa genome duplication is a general phenomenon through 10 chromosomes. The map includes five loci of FLC paralogues, which represent the previously reported BrFLC-1, -2, -3, and -5 and additionally identified BrFLC3 paralogues derived from local segmental duplication on R3.

We report the identification and characterization of the major repeats in the centromeric and peri-centromeric heterochromatin of Brassica rapa. The analysis involved the characterization of 88 629 bacterial artificial chromosomes (BAC) end sequences and the complete sequences of two BAC clones. We identified centromere-specific retrotransposons of Brassica (CRB) and various peri-centromere-specific retrotransposons (PCRBr). Three copies of the CRB were identified in one BAC clone as nested insertions within a tandem array of 24 copies of a 176 bp centromeric repeat, CentBr. A complex mosaic structure consisting of nine PCRBr elements and large blocks of 238 bp degenerate tandem repeats (TR238) were found in or near a derivative of 5S-25S rDNA sequences. The chromosomal positions of selected repeats were determined using in situ hybridization. These revealed that CRB is a major component of all centromeres in three diploid Brassica species and their allotetraploid relatives. However, CentBr was not detected in the most distantly related of the diploid species analyzed, B. nigra. PCRBr and TR238 were found to be major components in the pericentromeric heterochromatin blocks of four chromosomes of B. rapa. These repetitive elements were not identified in B. oleracea or B. nigra, indicating that they are A-genome-specific.