Genome-Wide Identification and Comparative Analysis of Conserved and Novel MicroRNAs in Grafted Watermelon by High-Throughput Sequencing

<div><p>MicroRNAs (miRNAs) are a class of endogenous small non-coding RNAs involved in the post-transcriptional gene regulation and play a critical role in plant growth, development and stresses response. However less is known about miRNAs involvement in grafting behaviors, especially with the watermelon (<i>Citrullus lanatus</i> L.) crop, which is one of the most important agricultural crops worldwide. Grafting method is commonly used in watermelon production in attempts to improve its adaptation to abiotic and biotic stresses, in particular to the soil-borne fusarium wilt disease. In this study, Solexa sequencing has been used to discover small RNA populations and compare miRNAs on genome-wide scale in watermelon grafting system. A total of 11,458,476, 11,614,094 and 9,339,089 raw reads representing 2,957,751, 2,880,328 and 2,964,990 unique sequences were obtained from the scions of self-grafted watermelon and watermelon grafted on-to bottle gourd and squash at two true-leaf stage, respectively. 39 known miRNAs belonging to 30 miRNA families and 80 novel miRNAs were identified in our small RNA dataset. Compared with self-grafted watermelon, 20 (5 known miRNA families and 15 novel miRNAs) and 47 (17 known miRNA families and 30 novel miRNAs) miRNAs were expressed significantly different in watermelon grafted on to bottle gourd and squash, respectively. MiRNAs expressed differentially when watermelon was grafted onto different rootstocks, suggesting that miRNAs might play an important role in diverse biological and metabolic processes in watermelon and grafting may possibly by changing miRNAs expressions to regulate plant growth and development as well as adaptation to stresses. The small RNA transcriptomes obtained in this study provided insights into molecular aspects of miRNA-mediated regulation in grafted watermelon. Obviously, this result would provide a basis for further unravelling the mechanism on how miRNAs information is exchanged between scion and rootstock in grafted watermelon, and its relevance to diverse biological processes and environmental adaptation.</p> </div

Comparison of expression patterns of miRNAs identified between Wm/Wm and Wm/BG (A) as well as Wm/Wm and Wm/Sq (B).

<p>Comparison of expression patterns of miRNAs identified between Wm/Wm and Wm/BG (A) as well as Wm/Wm and Wm/Sq (B).</p

Different expressed known miRNAs identified in watermelon grafted onto bottle gourd and squash rootstocks.

*<p>Log2 ratio of normalized miRNA expression in grafted watermelon compared with control; ↑and ↓:up- and down-regulated, respectively.</p

Sequence length distribution in Wm/Wm, Wm/BG and Wm/Sq.

<p>Sequence length distribution in Wm/Wm, Wm/BG and Wm/Sq.</p

Conserved miRNAS in watermelon.

<p>Y, yes; N, no.</p

Watermelon plants grafted onto watermelon (Wm/Wm) or onto bottle gourd (Wm/BG) and squash (Wm/Sq) at the two true-leaf stage.

<p>Watermelon plants grafted onto watermelon (Wm/Wm) or onto bottle gourd (Wm/BG) and squash (Wm/Sq) at the two true-leaf stage.</p

RNA secondary structure of the hairpin forming precursors of cla-miR21, cla-miR52, cla-miR72, cla-miR73 and cla-miR79.

<p>The putative mature miRNA sequences are shaded in green. Nucleotide positions are numbered starting from 5′ end of the precursor sequence.</p

The phenotypic analysis of root from MF and CMS of <i>Brassica juncea</i>.

<p>A, Root phenotype of MF, CMS and treated with 0.1 mmol/L and 0.5 mmol/L IAA. B, Statistic analysis of lateral root number. C, Statistic analysis of primary root length. Mean±SD values from 20 seedlings.</p

Average expression stability values (M) of the tested candidate reference genes determined by geNorm.

<p>Stability value of each reference gene was evaluated from four sample subsets: (A) scion leaves of different grafted watermelons submitted to N or P stress, (B) roots of grafted and non-grafted squash submitted to N or P stress, (C) roots of grafted and non-grafted bottle gourd submitted to N or P stress, (D) all scion leaves of different grafted watermelons and roots of grafted and non-grafted rootstocks under normal conditions. The most stable reference genes were measured during the stepwise exclusion of the least stable reference genes. The lower the M values the more stable expression of candidate reference genes.</p

The six structural types of RPS identified from strains of , , and that coaggregate with

<p><b>Copyright information:</b></p><p>Taken from "Streptococcal Receptor Polysaccharides: Recognition Molecules for Oral Biofilm Formation"</p><p>BMC Oral Health 2006;6(Suppl 1):S12-S12.</p><p>Published online 15 Jun 2006</p><p>PMCID:PMC2147599.</p><p></p> Partial O-acetylation of type 3G RPS is indicated. Lines indicate the location of GalNAcβ1-3Gal (Gn) or Galβ1-3GalNAc (G) recognition motifs within the hexa- or heptasaccharide repeating units of different RPS structural types