Heterosis and Combining Ability for Fruit Yield, Sweetness, β-Carotene, Ascorbic Acid, Firmness and Fusarium Wilt Resistance in Muskmelon (Cucumis melo L.) Involving Genetic Male Sterile Lines

Ten genetically diverse inbred lines, including two genic male sterile lines, of muskmelon (Cucumis melo L.) were crossed in a half-diallel to generate 45 F1 hybrids. These hybrids, along with the parental lines and commercial check, were evaluated for their fruit yield, level of phytochemicals and Fusarium wilt resistance. Both additive and non-additive genetic variances were important in governing the expression of all of the traits; however, the additive gene action for the fruit weight (g), flesh thickness (cm), rind thickness (mm), firmness (lb inch−2), β-carotene content (mg/100 g), non-additive variance for fruit yield (t ha−1), fruit number, total soluble solids (TSS, °Brix), ascorbic acid (mg/100 g) and reaction to Fusarium wilt were comparatively more important. The parental line MM-625 was the best general combiner for fruit yield, rind thickness and β-carotene content (mg/100 g). The exotic line Riogold was the best combiner for flesh thickness and firmness. The netted inbred line MM-610 was the best general combiner for fruit weight, ascorbic acid and reaction to Fusarium wilt. The inbred lines KP4HM-15 and MM-916 were the best general combiners for the number of fruits per vine and TSS. The best cross-combinations for fruit yield ha−1 and TSS were MS-1×M-610 and Kajri×MM-904, respectively. The hybrids KP4HM-15×MM Sel-103 and KP4HM-15×MM-1831 recorded the highest standard heterosis for fruit yield and TSS. The landrace-derived inbred lines Kajri, MM Sel-103 and KP4HM-15 produced moderate-to-highly FW-resistant hybrids. Out of the 121 SSR markers applied, 70 exhibited parental polymorphism. The markers DM0561, CMAAAGN14, TJ147, CMMS35_3, CMAGN45 and DE1337 identified specific/unique alleles in certain parental genotypes. Thus, the findings of this study revealed that the novel inbred lines can effectively be combined to generate heterotic F1 hybrids for yield and other traits, such as rind and flesh thickness, TSS, β-carotene content and firmness. Furthermore, SSR markers can potentially be utilized to confirm the genetic diversity among the parental lines, and for the DNA fingerprinting of F1 hybrids.

Phenotypic and molecular characterization of a set of tropical maize inbred lines from a public breeding program in Brazil

Background The characterization of genetic diversity and population differentiation for maize inbred lines from breeding programs is of great value in assisting breeders in maintaining and potentially increasing the rate of genetic gain. In our study, we characterized a set of 187 tropical maize inbred lines from the public breeding program of the Universidade Federal de Viçosa (UFV) in Brazil based on 18 agronomic traits and 3,083 single nucleotide polymorphisms (SNP) markers to evaluate whether this set of inbred lines represents a panel of tropical maize inbred lines for association mapping analysis and investigate the population structure and patterns of relationships among the inbred lines from UFV for better exploitation in our maize breeding program. Results Our results showed that there was large phenotypic and genotypic variation in the set of tropical maize inbred lines from the UFV maize breeding program. We also found high genetic diversity (GD = 0.34) and low pairwise kinship coefficients among the maize inbred lines (only approximately 4.00 % of the pairwise relative kinship was above 0.50) in the set of inbred lines. The LD decay distance over all ten chromosomes in the entire set of maize lines with r2 = 0.1 was 276,237 kb. Concerning the population structure, our results from the model-based STRUCTURE and principal component analysis methods distinguished the inbred lines into three subpopulations, with high consistency maintained between both results. Additionally, the clustering analysis based on phenotypic and molecular data grouped the inbred lines into 14 and 22 genetic divergence clusters, respectively. Conclusions Our results indicate that the set of tropical maize inbred lines from UFV maize breeding programs can comprise a panel of tropical maize inbred lines suitable for a genome-wide association study to dissect the variation of complex quantitative traits in maize, mainly in tropical environments. In addition, our results will be very useful for assisting us in the assignment of heterotic groups and the selection of the best parental combinations for new breeding crosses, mapping populations, mapping synthetic populations, guiding crosses that target highly heterotic and yielding hybrids, and predicting untested hybrids in the public breeding program UFV.

Integrated analysis of long non-coding RNAs and mRNAs reveals the regulatory network of maize seedling root responding to salt stress

Background Long non-coding RNAs (lncRNAs) play important roles in response to abiotic stresses in plants, by acting as cis- or trans-acting regulators of protein-coding genes. As a widely cultivated crop worldwide, maize is sensitive to salt stress particularly at the seedling stage. However, it is unclear how the expressions of protein-coding genes are affected by non-coding RNAs in maize responding to salt tolerance. Results The whole transcriptome sequencing was employed to investigate the differential lncRNAs and target transcripts responding to salt stress between two maize inbred lines with contrasting salt tolerance. We developed a flexible, user-friendly, and modular RNA analysis workflow, which facilitated the identification of lncRNAs and novel mRNAs from whole transcriptome data. Using the workflow, 12,817 lncRNAs and 8,320 novel mRNAs in maize seedling roots were identified and characterized. A total of 742 lncRNAs and 7,835 mRNAs were identified as salt stress-responsive transcripts. Moreover, we obtained 41 cis- and 81 trans-target mRNA for 88 of the lncRNAs. Among these target transcripts, 11 belonged to 7 transcription factor (TF) families including bHLH, C2H2, Hap3/NF-YB, HAS, MYB, WD40, and WRKY. The above 8,577 salt stress-responsive transcripts were further classified into 28 modules by weighted gene co-expression network analysis. In the salt-tolerant module, we constructed an interaction network containing 79 nodes and 3081 edges, which included 5 lncRNAs, 18 TFs and 56 functional transcripts (FTs). As a trans-acting regulator, the lncRNA MSTRG.8888.1 affected the expressions of some salt tolerance-relative FTs, including protein-serine/threonine phosphatase 2C and galactinol synthase 1, by regulating the expression of the bHLH TF. Conclusions The contrasting genetic backgrounds of the two inbred lines generated considerable variations in the expression abundance of lncRNAs and protein-coding transcripts. In the co-expression networks responding to salt stress, some TFs were targeted by the lncRNAs, which further regulated the salt tolerance-related functional transcripts. We constructed a regulatory pathway of maize seedlings to salt stress, which was mediated by the hub lncRNA MSTRG.8888.1 and participated by the bHLH TF and its downstream target transcripts. Future work will be focused on the functional revelation of the regulatory pathway.

The megabase-scale crossover landscape is independent of sequence divergence

Meiotic recombination frequency varies along chromosomes and strongly correlates with sequence divergence. However, the causality underlying this correlation is unclear. To untangle the relationship between recombination landscapes and polymorphisms, we characterized the genome-wide recombination landscape in the absence of polymorphisms, using Arabidopsis thaliana homozygous inbred lines in which a few hundred genetic markers were introduced through mutagenesis. We found that megabase-scale recombination landscapes in inbred lines are strikingly similar to the recombination landscapes in hybrids, with the sole exception of heterozygous large rearrangements where recombination is prevented locally. In addition, we found that the megabase-scale recombination landscape can be accurately predicted by chromatin features. Our results show that polymorphisms are not causal for the shape of the megabase-scale recombination landscape, rather, favor alternative models in which recombination and chromatin shape sequence divergence across the genome.

Recombinant inbred lines and next-generation sequencing enable rapid identification of candidate genes involved in morphological and agronomic traits in foxtail millet

We constructed recombinant inbred lines (RILs) between a Japanese and a Taiwanese landrace of foxtail millet and employed next-generation sequencing, such as flexible ddRAD-seq and Nanopore sequencing to identify the candidate genes involved in the crop evolution of foxtail millet. We successfully constructed a linkage map using flexible ddRAD-seq with parents and RILs and detected major QTLs for each of three traits: leaf sheath colors, spikelet-tipped bristles (stb), and days to heading (DTH). (1) For leaf sheath colors, we identified the C gene on chromosome IV. (2) We identified a homeobox (HOX14) gene for stb on chromosome II, which shows homology with HvVrs1 in barley. (3) Finally, we identified a QTL with a large effect on DTH on chromosome II. A parent of the RILs from Taiwan and Yugu1 had a Harbinger-like TE in intron 3 of this gene. We also investigated the geographical distribution of the TE insertion type of this gene and found that the insertion type is distributed in the northern part of East Asia and intensively in South and Southeast Asia, suggesting that loss/reduction of function of this gene plays an important role in spreading into the northern part of East Asia and subtropical and tropical zones.

Targeted Deletion of the First Intron of the Wxb Allele via CRISPR/Cas9 Significantly Increases Grain Amylose Content in Rice

Background The rice Waxy (Wx) gene plays a major role in seed amylose synthesis and consequently controls grain amylose content. Wx gene expression is highly regulated at the post-transcriptional level. In particular, the GT/TT polymorphism at the 5′splicing site of its 1st intron greatly affects this intron’s splicing efficiency and defines two predominant Wx alleles, Wxa and Wxb. Wxa rice often harbours intermediate to high amylose contents, whereas Wxb rice exhibits low to intermediate amylose contents. By deleting the Wx 1st intron using CRISPR/Cas9 technology, we generate a completely novel Wx allele and further investigate how intron removal affects Wx gene expression and rice grain amylose content. Results CRISPR/Cas9-mediated targeted deletion of the Wx 1st intron was performed on 4 rice inbred lines: KY131 (Wxb), X32 (Wxb), X35 (Wxa) and X55 (Wxlv). Deletion of the 1st intron occurred in 8.6–11.8% of the primary transformants of these 4 inbred lines. Compared to wild-type plants, amylose content was significantly increased from 13.0% to approximately 24.0% in KY131 and X32 mutant lines, which both carried the Wxb allele. However, no significant difference in amylose content was observed between wild-type plants and X35 and X55 mutant lines, which carried the Wxa and Wxlv alleles, respectively. Wx gene expression analysis of wild-type plants and mutants yielded results that were highly consistent with amylose content results. KY131 and X32 mutants accumulated increased levels of steady mRNA transcripts compared with wild-type plants, whereas steady mRNA levels were not altered in X35 and X55 mutants compared with wild-type plants. Grain quality, including appearance quality and eating and cooking quality, which are tightly associated with amylose content, was also assessed in wild-type and mutant plants, and data were presented and analysed. Conclusions This study presents a novel and rapid strategy to increase amylose content in inbred rice carrying a Wxb allele. Our data strongly suggest that the 1st intron of the Wx gene regulates Wx gene expression mainly at the post-transcriptional level in rice. This finding is in contrast to a previous hypothesis suggesting that it influences Wx gene transcription. In addition, removal of the first intron generates a completely novel Wx allele. Further studies on this new Wx allele will provide invaluable insights into the regulation of Wx gene expression, which will help researchers engineer new Wx alleles to facilitate the breeding of rice cultivars with better eating and cooking quality.

Effect of Meiotic Polyploidisation on Selected Morphological and Anatomical Traits in Interspecific Hybrids of Brassica oleracea × B. napus

In Brassica, interspecific hybridisation plays an important role in the formation of allopolyploid cultivars. In this study, the ploidy of F1 and F2 generations resulting from interspecific hybridisation between B. oleracea inbred lines of head cabbage (B. oleracea L. var. capitata) (2n = 18) and kale (B. oleracea L. var. acephala) (2n = 18) with inbred lines of rapeseed (B. napus L.) (2n = 38) was examined by flow cytometry analysis and chromosome observation. Furthermore, the effect of meiotic polyploidisation on selected phenotypic and anatomical traits was assessed. The F1 hybrids of head cabbage × rapeseed (S3) and kale × rapeseed crosses (S20) were allotriploids with 2n = 28 chromosomes, and nuclear DNA amounts of 1.97 (S3) and 1.99 pg (S20). These values were intermediate between B. oleracea and B. napus. In interspecific hybrids of the F2 generation, which were derived after self-pollination of F1 hybrids (FS3, FS20) or by open crosses between F1 generation hybrids (FC320, FC230), the chromosome numbers were similar 2n = 56 or 2n = 55, whereas the genome sizes varied between 3.81 (FS20) and 3.95 pg 2C (FC230). Allohexaploid F2 hybrids had many superior agronomic traits compared to parental B. napus and B. oleracea lines and triploid F1 hybrids. In the generative stage, they were characterised by larger flowers and flower elements, such as anthers and lateral nectaries. F2 hybrids were male and female fertile. The pollen viability of F2 hybrids was comparable to parental genotypes and varied from 75.38% (FS3) to 88.24% (FC320), whereas in triploids of F1 hybrids only 6.76% (S3) and 13.46% (S20) of pollen grains were fertile. Interspecific hybrids of the F2 generation derived by open crosses between plants of the F1 generation (FC320, FC230) had a better ability to set seed than F2 hybrids generated from the self-pollination of F1 hybrids. In the vegetative stage, F2 plants had bigger and thicker leaves, larger stomata, and significantly thicker layers of palisade and spongy mesophyll than triploids of the F1 generation and parental lines of B. oleracea and B. napus. The allohexaploid F2 hybrids analysed in this study can be used as innovative germplasm resources for further breeding new vegetable Brassica crops at the hexaploid level.

Evaluation of Drought Tolerance in Maize Inbred Lines Selected from the Shaan A Group and Shaan B Group

Drought is one of the most prevailing abiotic stresses affecting the growth, development, and productivity of maize. Knowledge of drought tolerance could help in maize improvement. However, less research has been done to comprehensively evaluate the drought tolerance of maize inbred lines. We used 27 elite maize inbred lines selected from Shaan A group and Shaan B group breeding populations to estimate their drought tolerance in 3 years 2 locations under normal field conditions and low irrigation. Using principal component analysis (PCA) and GGE biplots, all inbred lines, including the controls, could be divided into four types. Ten lines could be categorized as the high-yield drought-resistant type (‘KB081’, ‘KA105’, ‘KB417’, ‘KB215’, ‘KB-7’, ‘2013KB-37’, ‘KA203’, ‘2012KA-34’, ‘KA225’, and ‘91227’) because of their stability and wide adaptability. Compared with the controls, a large proportion of the inbred lines selected from Shaan A and Shaan B breeding populations demonstrated higher drought resistance. Our results suggest that multi-year drought screening can be used as a tool to improve the drought resistance of maize inbred lines and provide a scientific basis for making better use of the Shaan A and Shaan B maize inbred lines to breed new varieties and to identify existing drought-resistant maize varieties.