Morphogenesis of seed plants commences with highly stereotypical cell division sequences in early embryogenesis [1, 2]. Although a small number of transcription factors and a mitogen-activated protein (MAP) kinase cascade have been implicated in this process [3-8], pattern formation in early embryogenesis remains poorly understood. We show here that the Arabidopsis RKD4, a member of the RWP-RK motif-containing putative transcription factors [9], is required for this process. Loss-of-function rkd4 mutants were defective in zygotic cell elongation, as well as subsequent cell division patterns. As expected from this mutant phenotype, RKD4 was transcribed preferentially in early embryos. RKD4 possessed functional characteristics of transcription factors and was able to ectopically induce early embryo-specific genes when overexpressed in seedlings. Strikingly, induced overexpression of RKD4 primed somatic cells for embryogenesis independently of external growth regulators. These results reveal that RKD4 is a novel key regulator of the earliest stage of plant development.

SUMMARYActivation tagging is a powerful tool for discovering novel genes that are not easily identified by loss-offunction (lof) screening due to genetic redundancy or lethality. Although the current activation tagging system, which involves a viral enhancer sequence, has been used for a decade, alternative methods that allow organ-or tissue-specific activation are required to identify genes whose strong activation leads to loss of fertility or viability. Here, we established a GAL4/UAS activation-tagging system in Arabidopsis thaliana. Host plants that express a synthetic transcription activator GAL4:VP16 (GV) in an organ-or tissue-specific manner were transformed with a T-DNA harboring tandem copies of UAS, a GAL4-binding sequence. Using a post-embryonic and root-specific GV-expressing line as the host plant, we isolated several dominant mutants with abnormal root tissue patterns, designated as uas-tagged root patterning (urp) mutants, and identified their causal genes. Notably, most URP genes encoded putative transcription factors, indicating that the GAL4/UAS activation tagging system effectively identifies genes with regulatory functions. lof phenotypes of most URP genes were either local patterning defects or visible only if homologous genes were disrupted simultaneously or independently. Systemic overexpression of some URP genes resulted in seedling lethality. These results indicate that GAL4/UAS activation tagging is a powerful method for identifying genes with biological functions that are not readily identified by conventional screening methods.

Double flower and hortensia (mophead) hydrangea (Hydrangea macrophylla (Thunb.) Ser.) traits are recessively inherited. Cross breeding of these traits in hydrangea is difficult because it takes about two years from crossing to flowering. In this study, we aimed to obtain DNA linkage markers that would allow accelerated selection of these traits. We used next-generation sequencing to comprehensively collect DNA sequences from the 'Kirakiraboshi' with a double flower and lacecap inflorescence and the 'Frau Yoshimi' with a single flower and hortensia inflorescence, and designed simple sequence repeat (SSR) primer pairs for map construction. We screened 768 SSR primer pairs in 93 F 2 progeny derived from 'Kirakiraboshi' and 'Frau Yoshimi'. We identified 147 loci, which were expanded to 18 linkage groups with a total map length of 980 cM. Linkage analysis identified that both the double flower trait from 'Kirakiraboshi' (d Kira ) and the hortensia trait from 'Frau Yoshimi' (h Frau ) were located on linkage group KF_4. Detailed linkage analysis using 351 F 2 progeny revealed a 34.8 cM map length between the two loci and identified two tightly linked SSR markers, STAB045 for d Kira and HS071 for h Frau . Genetic analysis suggested that double flower and hortensia traits are each controlled by a single recessive gene. Together, the linkage map, SSR markers, and genetic information obtained in this study will be useful for future hydrangea breeding.

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Abstract:Retrotransposons have been used frequently for the development of 23 molecular markers by using their insertion polymorphisms among cultivars, because 24 multiple copies of these elements are dispersed throughout the genome and inserted 25 copies are inherited genetically. Although a large number of long terminal repeat 26 (LTR) retrotransposon families exist in the higher eukaryotic genomes, the 27 identification of families that show high insertion polymorphism has been challenging. 28Here, we performed an efficient screening of these retrotransposon families using an 29Illumina HiSeq2000 sequencing platform with comprehensive LTR library 30 construction based on the primer binding site (PBS), which is located adjacent to the 5 31 LTR and has a motif that is universal and conserved among LTR retrotransposon 32 families. The paired-end sequencing library of the fragments containing a large number 33 of LTR sequences and their insertion sites was sequenced for seven strawberry 34 cultivars (Fragaria x ananassa Duchesne) and one diploid wild species (F. vesca). 35Among them, we screened 24 families with a "unique" insertion site that appeared only 36 in one variety and not in any others, assuming that this type of insertion should have 37 occurred quite recently. Finally, we confirmed experimentally the selected LTR 38 families showed high insertion polymorphisms among closely related cultivars. 39 40