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Repetitive sequences constitute a significant component of most eukaryotic genomes, and the isolation and characterization of repetitive DNA sequences provide an insight into the organization of the genome of interest. Here, we report the isolation and molecular analysis of a novel tandemly organized repetitive DNA sequence from the genome of Citrus limon. Digestion of C. limon DNA with Hinf I produced a prominent fragment of approximately 300 bp. Southern blotting revealed a ladder composed of DNA fragments that were multimers of the 300-bp Hinf I band. Thus, Hinf I digestion revealed a novel satellite, which we have called the C. limon satellite DNA 300 (CL300). Sequence analysis shows significant homology between a portion of the CL300 monomer and the transposase box of an En/Spm-like element. The CL300 satellite was also detected in grapefruit, sour orange, trifoliate orange and kumquat. These results suggest that the CL300 repeat is an ancient satellite, and we propose that a significant portion originated by amplification of a genomic region containing the En/Spm-like transposase element.
The plant nuclear genome is largely composed of mobile DNA, which can rearrange genomes and other individual gene structure and also affect gene regulation through various promoted activities: transposition, insertion, excision, chromosome breakage, and ectopic recombination. Ty1-copia-like retrotransposon is a widespread class of transposable elements in the plant kingdom, representing a large part of the total DNA content. Here, a novel retrotransposon-like sequence was isolated and identified as the Ty1-copia-like reverse transcriptase domain (named here CLCoy1), based on the homology of known elements. Fluorescence in situ hybridization, revealed that CLCoy1 was mainly located in telomeric and sub-telomeric regions along the Citrus chromosomes. CLCoy1 composes 3.6% of the genome and, interestingly, while transposons are mostly specific to a species, this element was identified in other Citrus species such as Citrus aurantium, Fortunella margarita and Citrus paradisi, but undetected in Poncirus trifoliata. We also determined that wounding, salt and cell culture stress produced transcriptional activation of this novel retroelement in Citrus limon. The novel Ty1-copia-like element CLCoy1 may have played a major role in shaping genome structure and size during Citrus species evolution.
In this work, a seed-set-based screening was performed on 70 lines of Arabidopsis thaliana after activation tagging mutagenesis to identify mutations in reproductive mechanisms. Five mutants showed significantly lower seed set than the wild type and confirmed the phenotype in the progeny. This phenotype was linked with the marker gene bar carried by T-DNA conferring glufosinate resistance. Genetic analysis revealed that the mutation inheritance was sporophytic in 3 mutants and gametophytic in 2 mutants. In addition, 2 mutants had an extra T-DNA copy. Thus activation tagging can be an effective strategy to identify new mutations affecting sporogenesis or gametogenesis.
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