Fine mapping and discovery of candidate genes for seed size in watermelon by genome survey sequencing.

Fine mapping and discovery of candidate genes underlying seed size are important for modern watermelon breeding. Here, by using a high-resolution genetic map and whole-genome genetic variation detection aided by genome survey sequencing, we fine mapped and discovered candidate genes for seed size in watermelon. QTL (quantitative trait locus) mapping identified two pleiotropic QTLs for seed size, namely, qSS4 and qSS6, using a high-density genetic map constructed by specific length amplified fragment sequencing. qSS6 explained 93.00%, 94.11% and 95.26% of the phenotypic variation in thousand-seed weight, seed length and seed width, respectively, and was defined as a major QTL. Then, high-coverage re-sequencing of two parental lines detected a total of 193,395 SNPs (single nucleotide polymorphisms) and 45,065 indels (insertions/deletions), which corresponded to a frequency of 534 SNPs/Mb and 124 indels/Mb. Based on the genetic variation in the two parental lines, newly developed PCR-based markers allowed the region of qSS6 to be narrowed to 55.5 kb. Three potential candidates were identified, including a known seed size regulator in rice, SRS3. Taken together, our results reveal successful rapid fine mapping and discovery of candidate genes for seed size in watermelon, which could be applied to many traits of interest in plants.

Genetic mapping of the LOBED LEAF 1 (ClLL1) gene to a 127.6-kb region in watermelon (Citrullus lanatus L.).

The lobed leaf character is a unique morphologic trait in crops, featuring many potential advantages for agricultural productivity. Although the majority of watermelon varieties feature lobed leaves, the genetic factors responsible for lobed leaf formation remain elusive. The F2:3 leaf shape segregating population offers the opportunity to study the underlying mechanism of lobed leaf formation in watermelon. Genetic analysis revealed that a single dominant allele (designated ClLL1) controlled the lobed leaf trait. A large-sized F3:4 population derived from F2:3 individuals was used to map ClLL1. A total of 5,966 reliable SNPs and indels were identified genome-wide via a combination of BSA and RNA-seq. Using the validated SNP and indel markers, the location of ClLL1 was narrowed down to a 127.6-kb region between markers W08314 and W07061, containing 23 putative ORFs. Expression analysis via qRT-PCR revealed differential expression patterns (fold-changes above 2-fold or below 0.5-fold) of three ORFs (ORF3, ORF11, and ORF18) between lobed and non-lobed leaf plants. Based on gene annotation and expression analysis, ORF18 (encoding an uncharacterized protein) and ORF22 (encoding a homeobox-leucine zipper-like protein) were considered as most likely candidate genes. Furthermore, sequence analysis revealed no polymorphisms in cDNA sequences of ORF18; however, two notable deletions were identified in ORF22. This study is the first report to map a leaf shape gene in watermelon and will facilitate cloning and functional characterization of ClLL1 in future studies.

The Ethylene Biosynthesis Gene CitACS4 Regulates Monoecy/Andromonoecy in Watermelon (Citrullus lanatus).

Monoecious and andromonoecious cultivars of watermelon are characterised by the production of male and female flower or male and hermaphrodite flowers, respectively. The segregation analysis in the offspring of crosses between monoecious and andromonoecious lines has demonstrated that this trait is controlled by a single gene pair, being the monoecious allele M semi-dominant to the andromonoecious allele A. The two studied F1 hybrids (MA) had a predominantly monoecious phenotype since both produced not only female flowers, but also bisexual flowers with incomplete stamens, and hermaphrodite flowers with pollen. Given that in other cucurbit species andromonoecy is conferred by mutations in the ethylene biosynthesis genes CmACS7, CsACS2 and CpACS27A we have cloned and characterised CitACS4, the watermelon gene showing the highest similarity with the formers. CitACS4 encoded for a type ACS type III enzyme that is predominantly expressed in pistillate flowers of watermelon. In the andromonoecious line we have detected a missense mutation in a very conserved residue of CitACS4 (C364W) that cosegregates with the andromonoecious phenotype in two independent F2 populations, concomitantly with a reduction in ethylene production in the floral buds that will develop as hermaphrodite flowers. The gene does not however co-segregates with other sex expression traits regulated by ethylene in this species, including pistillate flowering transition and the number of pistillate flowers per plant. These data indicate that CitAC4 is likely to be involved in the biosynthesis of the ethylene required for stamen arrest during the development of female flowers. The C364W mutation would reduce the production of ethylene in pistillate floral buds, promoting the conversion of female into hermaphrodite flowers, and therefore of monoecy into andromonoecy.

A non-destructive genotyping system from a single seed for marker-assisted selection in watermelon.

Genomic tools for watermelon breeding are becoming increasingly available. A high throughput genotyping system would facilitate the use of DNA markers in marker-assisted selection. DNA extraction from leaf material requires prior seed germination and is often time-consuming and cost prohibitive. In an effort to develop a more efficient system, watermelon seeds of several genotypes and various seed sizes were sampled by removing ⅓ or ½ sections from the distal ends for DNA extraction, while germinating the remaining proximal parts of the seed. Removing ⅓ of the seed from the distal end had no effect on seed germination percentage or seedling vigor. Different DNA extraction protocols were tested to identify a method that could yield DNA of sufficient quality for amplification by polymerase chain reaction. A sodium dodecyl sulfate extraction protocol with 1% polyvinylpyrrolidone yielded DNA that could be amplified with microsatellite primers and was free of pericarp contamination. In this study, an efficient, non-destructive genotyping protocol for watermelon seed was developed.

[Pharmacognostical studies on imported and cultivated Citrullus colocynthis].

OBJECTIVE: In order to use Citrullus colocynthis safety and correctly,The pharmacognosy of imported and cultivated Citrullus colocynthis was systematically studied. METHODS: The morphological characters, microscopical identification, and thin layer Chromatograpy (TLC) were studied. RESULTS: The characters of import and cultivated Citrullus colocynthis were approximately identical, but there were still some differences between them, such as: the fruit size of cultivated Citrullus colocynthis was largish, granulose were more, trachea were thicker, collenchyma were longer, the colour of spots in TCL were more clear. CONCLUSION: There were seldon difference between import and cultivated Citrullus colocynthis, and cultivated Citrullus colocynthis could take the place of import's if planted on a large scale.

Divergent metabolome and proteome suggest functional independence of dual phloem transport systems in cucurbits.

Cucurbitaceous plants (cucurbits) have long been preferred models for studying phloem physiology. However, these species are unusual in that they possess two different phloem systems, one within the main vascular bundles [fascicular phloem (FP)] and another peripheral to the vascular bundles and scattered through stem and petiole cortex tissues [extrafascicular phloem (EFP)]. We have revisited the assumption that the sap released after shoot incision originates from the FP, and also investigated the long-standing question of why the sugar content of this sap is ~30-fold less than predicted for requirements of photosynthate delivery. Video microscopy and phloem labeling experiments unexpectedly reveal that FP very quickly becomes blocked upon cutting, whereas the extrafascicular phloem bleeds for extended periods. Thus, all cucurbit phloem sap studies to date have reported metabolite, protein, and RNA composition and transport in the relatively minor extrafascicular sieve tubes. Using tissue dissection and direct sampling of sieve tube contents, we show that FP in fact does contain up to 1 M sugars, in contrast to low-millimolar levels in the EFP. Moreover, major phloem proteins in sieve tubes of FP differ from those that predominate in the extrafascicular sap, and include several previously uncharacterized proteins with little or no homology to databases. The overall compositional differences of the two phloem systems strongly indicate functional isolation. On this basis, we propose that the fascicular phloem is largely responsible for sugar transport, whereas the extrafascicular phloem may function in signaling, defense, and transport of other metabolites.

The antioxidant activity and polyphenolic contents of different plant seeds extracts.

Different plant seeds extracts of Citrus sinensis, Hordeum sativum, Triticum sativum, Canna indica, Citrullus vulgaris and Capsicum annuum were evaluated for their antioxidant activity by the following methods: 2,2-diphenyl-1-pycril-hydrazyl (DPPH) radical scavenging, reducing power, RBCs hemolysis and linoleic acid oxidation, a long with the determination of total phenolic and flavonoids contents. All the methanolic extracts showed high antioxidant activity and have high contents of phenolic and flavonoid. The Canna indica extract exhibited strong antioxidant as a reducing power and as DPPH radical-scavenging (3.61 absorbance, 87.12%, respectively), while the Hordeum sativum extract exhibited highest inhibitory effect on RBCs hemolysis (59.55%) and the Capsicum annuum extract has highest inhibitory effect on linoleic acid peroxidation (65.06%).

Qualitative inheritance of rind pattern and flesh color in watermelon.

Watermelon [Citrullus lanatus (Thunb.) Matsum. & Nakai var. lanatus] is a diverse species, with fruits of different sizes, shapes, rind patterns, and flesh colors. This study measured the inheritance of novel rind phenotypes and verified the genetics of white, red, salmon yellow, and canary yellow flesh colors. For each of the 11 crosses, six generations (P(a)S1, P(b)S1, F1, F2, BC1P(a), and BC1P(b)) were produced to form 11 families. Three new genes were identified and designated as follows: Scr for the scarlet red flesh color of Dixielee and Red-N-Sweet, Yb for the yellow belly (ground spot) of Black Diamond Yellow Belly, and ins for the intermittent stripes of Navajo Sweet. The inheritance of the C gene for the canary yellow flesh color was verified as single dominant, and a new inbred type line was developed possessing that gene. Aberrations in the segregation of red, white, and salmon yellow flesh colors were recorded, raising questions on the inheritance of these traits. Finally, the spotted phenotype from Moon and Stars was combined with light green and gray rind patterns for the development of novel cultivars with distinctive rind patterns.

Inheritance of egusi seed type in watermelon.

An unusual seed mutant in watermelon (Citrullus lanatus var. lanatus) has seeds with a fleshy pericarp, commonly called egusi seeds. The origin of the phenotype is unknown, but it is widely cultivated in Nigeria for the high protein and carbohydrate content of the edible seeds. Egusi seeds have a thick, fleshy pericarp that appears during the second to third week of fruit development. We studied the inheritance of this phenotype in crosses of normal seeded Charleston Gray and Calhoun Gray with two plant introduction accessions, PI 490383w and PI 560006, having the egusi seed type. We found that the egusi seed type is controlled by a single recessive gene, and the symbol eg was assigned.

[Histological studies on rinds of 18 species of Gualou].

OBJECTIVE: To make clear the histological characteristics of rinds for eighteen species of Gualou. METHOD: Observing the cross sections and separate divide of the rinds with light microscope. RESULT AND CONCLUSION: Of eighteen rinds of Gualou, each trichome on exocarp, wood fiber and thickness of ring of stone cell are different from one another, this distinction can be used to identify the different rinds of Gualou.