Characterization of the OmyY1 Region on the Rainbow Trout Y Chromosome

We characterized the male-specific region on the Y chromosome of rainbow trout, which contains both sdY (the sex-determining gene) and the male-specific genetic marker, OmyY1. Several clones containing the OmyY1 marker were screened from a BAC library from a YY clonal line and found to be part of an 800 kb BAC contig. Using fluorescencein situhybridization (FISH), these clones were localized to the end of the short arm of the Y chromosome in rainbow trout, with an additional signal on the end of the X chromosome in many cells. We sequenced a minimum tiling path of these clones using Illumina and 454 pyrosequencing. The region is rich in transposons and rDNA, but also appears to contain several single-copy protein-coding genes. Most of these genes are also found on the X chromosome; and in several cases sex-specific SNPs in these genes were identified between the male (YY) and female (XX) homozygous clonal lines. Additional genes were identified by hybridization of the BACs to the cGRASP salmonid 4x44K oligo microarray. By BLASTn evaluations using hypothetical transcripts of OmyY1-linked candidate genes as query against several EST databases, we conclude at least 12 of these candidate genes are likely functional, and expressed.

Mapping of Micro-Tom BAC-End Sequences to the Reference Tomato Genome Reveals Possible Genome Rearrangements and Polymorphisms

A total of 93,682 BAC-end sequences (BESs) were generated from a dwarf model tomato, cv. Micro-Tom. After removing repetitive sequences, the BESs were similarity searched against the reference tomato genome of a standard cultivar, “Heinz 1706.” By referring to the “Heinz 1706” physical map and by eliminating redundant or nonsignificant hits, 28,804 “unique pair ends” and 8,263 “unique ends” were selected to construct hypothetical BAC contigs. The total physical length of the BAC contigs was 495, 833, 423 bp, covering 65.3% of the entire genome. The average coverage of euchromatin and heterochromatin was 58.9% and 67.3%, respectively. From this analysis, two possible genome rearrangements were identified: one in chromosome 2 (inversion) and the other in chromosome 3 (inversion and translocation). Polymorphisms (SNPs and Indels) between the two cultivars were identified from the BLAST alignments. As a result, 171,792 polymorphisms were mapped on 12 chromosomes. Among these, 30,930 polymorphisms were found in euchromatin (1 per 3,565 bp) and 140,862 were found in heterochromatin (1 per 2,737 bp). The average polymorphism density in the genome was 1 polymorphism per 2,886 bp. To facilitate the use of these data in Micro-Tom research, the BAC contig and polymorphism information are available in the TOMATOMICS database.

The Rvi15 (Vr2) Apple Scab Resistance Locus Contains Three TIR-NBS-LRR Genes

Scab caused by the pathogen Venturia inaequalis is considered the most important fungal disease of cultivated apple (Malus × domestica Borkh.). In all, 16 monogenic resistances against scab have been found in different Malus spp. and some of them are currently used in apple breeding for scab-resistant cultivars. However, the self incompatibility and the long generation time of Malus spp. together with the high standards of fruit quality demanded from the fresh market render the breeding of high-quality cultivars in apple a long and expensive task. Therefore, the cloning of disease resistance genes and the use of the cloned genes for the transformation of high-quality apple cultivars could be an approach to solve these drawbacks. We report the construction of a bacterial artificial chromosome (BAC) contig spanning the Rvi15 (Vr2) apple scab resistance locus using two GMAL 2473 BAC libraries. A single BAC clone of the contig was sufficient to span the resistance locus. The BAC clone was completely sequenced, allowing identification of a sequence of 48.6 kb going from the two closest markers (ARGH17 and 77G20RP) bracketing Rvi15 (Vr2). Analysis of the 48.6-kb sequence revealed the presence of three putative genes characterized by a Toll and mammalian interleukin-1 receptor protein nucleotide-binding site leucine-rich repeat structure. All three genes were found to be transcribed.

Identification by FISH of 4 novel partner loci of PRDM16 in myeloid malignancies

11037 Background: PRDM16 is a gene located on 1p36.32 that encodes for a zinc finger transcription factor and contains an N-terminal PR domain. It has been shown to be involved in the reciprocal translocation t(1;3)(p36;q21) and more rarely the t(1;21)(p36;q22) which both occur in myelodysplastic syndromes (MDS) and acute myeloid leukemias (AML). These translocations result in the overexpression of a truncated version of the PRDM16 protein that lacks the PR domain. This overexpression might play an important role in the pathogenesis of MDS and AML in blocking myeloid differentiation. This raises the question whether PRDM16 is rearranged with other partner genes and whether its sequences play the most important role in the oncogenic process attributed to those translocations. Methods: We studied 35 myeloid malignancies, 12 lymphoid malignancies and 3 undifferentiated acute leukemias with 1p36 abnormalities by fluorescent in situ hybridization (FISH) with a bacterial artificial chromosomes (BAC) contig containing 50 BAC probes on 1p36. Results: In addition to the known t(1;3)(p36;q21) (11 cases) and t(1;21)(p36;q22) (1 case) involving RPN1 andAML1/RUNX1 respectively in myeloid malignancies, we specifically found PRDM16 to be rearranged in 4 additional translocations : a t(1;12)(p36;p13) in an AML-M4, a t(1;7)(p36;p12) in a MDS, an add(1)(p36) in an AML-M2 and a t(1;2)(p36;p12) in a relapsed AML-M4. We identified the respective candidate partner loci : TEL/ETV6, IKZF1, CDH4 and a non-coding unknown sequence. Conclusions: In our series of 50 cases of hematological malignancies with 1p36 abnormalities, PRDM16 was involved in about 45% of myeloid malignancies, and was never involved in lymphoid malignancies. PRDM16 is supposed to have similar oncogenic properties as MDS1/EVI-1(3q26), another gene encoding for a zinc finger protein and acting as a transcriptional regulatory factor with 2 isoforms. Interestingly, the shortest isoform of MDS/EVI-1, lacking the PR domain, is supposed to have an oncogenic effect due to its translocation-induced upregulation in AML. Further characterization of these new partner genes and functional studies should give us more insight into the pathogenesis of AML and MDS mediated by PRDM16, and the role of its partner genes. No significant financial relationships to disclose.

A one-megabase physical map provides insights on gene organization in the enormous mitochondrial genome of cucumber

Cucumber ( Cucumis sativus ) has one of the largest mitochondrial genomes known among all eukaryotes, due in part to the accumulation of short 20 to 60 bp repetitive DNA motifs. Recombination among these repetitive DNAs produces rearrangements affecting organization and expression of mitochondrial genes. To more efficiently identify rearrangements in the cucumber mitochondrial DNA, we built two nonoverlapping 800 and 220 kb BAC contigs and assigned major mitochondrial genes to these BACs. Polymorphism carried on the largest BAC contig was used to confirm paternal transmission. Mitochondrial genes were distributed across BACs and physically distant, although occasional clustering was observed. Introns in the nad1, nad4, and nad7 genes were larger than those reported in other plants, due in part to accumulation of short repetitive DNAs and indicating that increased intron sizes contributed to mitochondrial genome expansion in cucumber. Mitochondrial genes atp6 and atp9 are physically close to each other and cotranscribed. These physical contigs will be useful for eventual sequencing of the cucumber mitochondrial DNA, which can be exploited to more efficiently screen for unique rearrangements affecting mitochondrial gene expression.