ALBERT

Description: Japanese flounder ( Paralichthys olivaceus ) is one of the economic important fish in China. Sexual dimorphism, especially the different growth rates and body sizes between two sexes, makes this fish a good model to investigate mechanisms responsible for such dimorphism for both fundamental questions in evolution and applied topics in aquaculture. However, the lack of "omics" data has hindered the process. The recent advent of RNA-sequencing technology provides a robust tool to further study characteristics of genomes of nonmodel species. Here, we performed de novo transcriptome sequencing for a double haploid Japanese flounder individual using Illumina sequencing. A single lane of paired-end sequencing produced more than 27 million reads. These reads were assembled into 107,318 nonredundant transcripts, half of which (51,563; 48.1%) were annotated by blastx to public protein database. A total of 1051 genes that had potential alternative splicings were detected by Chrysalis implemented in Trinity software. Four of 10 randomly picked genes were verified truly containing alternative splicing by cloning and Sanger sequencing. Notably, using a doubled haploid Japanese flounder individual allow us to analyze gene duplicates. In total, 3940 "single-nucleotide polymorphisms" were detected form 1859 genes, which may have happened gene duplicates. This study lays the foundation for structural and functional genomics studies in Japanese flounder.

Description: The internode length above the uppermost ear (ILAU) is an important influencing factor for canopy architecture in maize. Analyzing the genetic characteristics of internode length is critical for improving plant population structure and increasing photosynthetic efficiency. However, the genetic control of ILAU has not been determined. In this study, quantitative trait loci (QTL) for internode length at five positions above the uppermost ear were identified using four sets of recombinant inbred line (RIL) populations in three environments. Genetic maps and initial QTL were integrated using meta-analyses across the four populations. Seventy QTL were identified: 16 in population 1; 14 in population 2; 25 in population 3; and 15 in population 4. Individual effects ranged from 5.36% to 26.85% of phenotypic variation, with 27 QTL 〉10%. In addition, the following common QTL were identified across two populations: one common QTL for the internode length of all five positions; one common QTL for the internode length of three positions; and one common QTL for the internode length of one position. In addition, four common QTL for the internode length of four positions were identified in one population. The results indicated that the ILAU at different positions above the uppermost ear could be affected by one or several of the same QTL. The traits may also be regulated by many different QTL. Of the 70 initial QTL, 46 were integrated in 14 meta-QTL (mQTLs) by meta-analysis, and 17 of the 27 initial QTL with R 2  〉10% were integrated in 7 mQTLs. Four of the key mQTLs (mQTL2-2, mQTL3-2, mQTL5-1, mQTL5-2, and mQTL9) in which the initial QTL displayed R 2 〉10% included four to 11 initial QTL for an internode length of four to five positions from one or two populations. These results may provide useful information for marker-assisted selection to improve canopy architecture.

Description: OsWUS has recently been shown to be a transcription factor gene critical for tiller development and fertility in rice. The OsWUS protein consists of three conserved structural domains, but their biological functions are still unclear. We discovered a new rice mutant resulting from tissue culture, which hardly produced tillers and exhibited complete female sterility. The male and female floral organs of the mutant were morphologically indistinguishable from those of the wild type. We named the mutant srt1 for completely s terile and r educed t illering 1. Map-based cloning revealed that the mutant phenotypes were caused by a mutation in OsWUS . Compared with the two previously reported null allelic mutants of OsWUS ( tab1-1 and moc3-1 ), which could produce partial N-terminal peptides of OsWUS, the srt1 protein contained a deletion of only seven amino acids within the conserved homeobox domain of OsWUS. However, the mutant phenotypes (monoculm and female sterility) displayed in srt1 were as typical and severe as those in tab1-1 and moc3-1 . This indicates that the homeobox domain of SRT1 is essential for the regulation of tillering and sterility in rice. In addition, srt1 showed an opposite effect on panicle development to that of the two null allelic mutants, implying that the srt1 protein might still have partial or even new functions on panicle development. The results of this study suggest that the homeobox domain is pivotal for OsWUS function.

Description: The fat-1 gene from Caenorhabditis elegans encodes a fatty acid desaturase which was widely studied due to its beneficial function of converting n-6 polyunsaturated fatty acids (n-6PUFAs) to n-3 polyunsaturated fatty acids (n-3PUFAs). To date, many fat-1 transgenic animals have been generated to study disease pathogenesis or improve meat quality. However, all of them were generated using a random integration method with variable transgene expression levels and the introduction of selectable marker genes often raise biosafety concern. To this end, we aimed to generate marker-free fat-1 transgenic pigs in a site-specific manner. The Rosa26 locus, first found in mouse embryonic stem cells, has become one of the most common sites for inserting transgenes due to its safe and ubiquitous expression. In our study, the fat-1 gene was inserted into porcine Rosa 26 (pRosa26) locus via Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/CRISPR-associated 9 (Cas9) system. The Southern blot analysis of our knock-in pigs indicated a single copy of the fat-1 gene at the pRosa26 locus. Furthermore, this single-copy fat-1 gene supported satisfactory expression in a variety of tissues in F1 generation pigs. Importantly, the gas chromatography analysis indicated that these fat-1 knock-in pigs exhibited a significant increase in the level of n-3PUFAs, leading to an obvious decrease in the n-6PUFAs/n-3PUFAs ratio from 9.36 to 2.12 (*** P 〈 0.0001). Altogether, our fat-1 knock-in pigs hold great promise for improving the nutritional value of pork and serving as an animal model to investigate therapeutic effects of n-3PUFAs on various diseases.

Description: The Dongxiang Blue-shelled chicken is one of the most valuable Chinese indigenous poultry breeds. However, compared to the Italian native White Leghorn, although this Chinese breed possesses numerous favorable characteristics, it also exhibits lower growth performance and fertility. Here, we utilized genotyping sequencing data obtained via genome reduction on a sequencing platform to detect 100,114 single nucleotide polymorphisms and perform further biological analysis and functional annotation. We employed cross-population extended haplotype homozygosity, eigenvector decomposition combined with genome-wide association studies (EigenGWAS), and efficient mixed-model association expedited methods to detect areas of the genome that are potential selected regions (PSR) in both chicken breeds, and performed gene ontology (GO) enrichment and quantitative trait loci (QTL) analyses annotating using the Kyoto Encyclopedia of Genes and Genomes. The results of this study revealed a total of 2424 outlier loci ( p -value 〈0.01), of which 2144 occur in the White Leghorn breed and 280 occur in the Dongxiang Blue-shelled chicken. These correspond to 327 and 94 PSRs containing 297 and 54 genes, respectively. The most significantly selected genes in Blue-shelled chicken are TMEM141 and CLIC3 , while the SLCO1B3 gene, related to eggshell color, was identified via EigenGWAS. We show that the White Leghorn genes JARID2 , RBMS3 , GPC3 , TRIB2 , ROBO1 , SAMSN1 , OSBP2 , and IGFALS are involved in immunity, reproduction, and growth, and thus might represent footprints of the selection process. In contrast, we identified six significantly enriched pathways in the Dongxiang Blue-shelled chicken that are related to amino acid and lipid metabolism as well as signal transduction. Our results also reveal the presence of a GO term associated with cell metabolism that occurs mainly in the White Leghorn breed, while the most significant QTL regions mapped to the Chicken QTL Database (GG_4.0) for the Dongxiang Blue-shelled breed are predominantly related to lesions, bone mineral content, and other related traits compared to tibia length and body weight ( i.e. , at 14, 28, 42, and 70 d) in the White Leghorn. The results of this study highlight differences in growth, immunity, and egg quality traits between the two breeds, and provide a foundation for the exploration of their genetic mechanisms.

Description: Rapid and highly efficient mating-type switching of Saccharomyces cerevisiae enables a wide variety of genetic manipulations, such as the construction of strains, for instance, isogenic haploid pairs of both mating-types, diploids and polyploids. We used the CRISPR/Cas9 system to generate a double-strand break at the MAT locus and, in a single cotransformation, both haploid and diploid cells were switched to the specified mating-type at ~80% efficiency. The mating-type of strains carrying either rod or ring chromosome III were switched, including those lacking HML α and HMR a cryptic mating loci. Furthermore, we transplanted the synthetic yeast chromosome V to build a haploid polysynthetic chromosome strain by using this method together with an endoreduplication intercross strategy. The CRISPR/Cas9 mating-type switching method will be useful in building the complete synthetic yeast (Sc2.0) genome. Importantly, it is a generally useful method to build polyploids of a defined genotype and generally expedites strain construction, for example, in the construction of fully a/a /α/α isogenic tetraploids.

Description: Following the two rounds of whole-genome duplication that occurred during deuterostome evolution, a third genome duplication event occurred in the stem lineage of ray-finned fishes. This teleost-specific genome duplication is thought to be responsible for the biological diversification of ray-finned fishes. DEAD-box polypeptide 3 (DDX3) belongs to the DEAD-box RNA helicase family. Although their functions in humans have been well studied, limited information is available regarding their function in teleosts. In this study, two teleost Ddx3 genes were first identified in the transcriptome of Japanese flounder ( Paralichthys olivaceus ). We confirmed that the two genes originated from teleost-specific genome duplication through synteny and phylogenetic analysis. Additionally, comparative analysis of genome structure, molecular evolution rate, and expression pattern of the two genes in Japanese flounder revealed evidence of subfunctionalization of the duplicated Ddx3 genes in teleosts. Thus, the results of this study reveal novel insights into the evolution of the teleost Ddx3 genes and constitute important groundwork for further research on this gene family.

Description: We constructed a very-high-density, whole-genome marker map (WGMM) for cotton by using 18,597 DNA markers corresponding to 48,958 loci that were aligned to both a consensus genetic map and a reference genome sequence. The WGMM has a density of one locus per 15.6 kb, or an average of 1.3 loci per gene. The WGMM was anchored by the use of colinear markers to a detailed genetic map, providing recombinational information. Mapped markers occurred at relatively greater physical densities in distal chromosomal regions and lower physical densities in the central regions, with all 1 Mb bins having at least nine markers. Hotspots for quantitative trait loci and resistance gene analog clusters were aligned to the map and DNA markers identified for targeting of these regions of high practical importance. Based on the cotton D genome reference sequence, the locations of chromosome structural rearrangements plotted on the map facilitate its translation to other Gossypium genome types. The WGMM is a versatile genetic map for marker assisted breeding, fine mapping and cloning of genes and quantitative trait loci, developing new genetic markers and maps, genome-wide association mapping, and genome evolution studies.

Description: Black or dark brown (phaeoid) fungi cause cutaneous, subcutaneous, and systemic infections in humans. Black fungi thrive in stressful conditions such as intense light, high radiation, and very low pH. Wangiella ( Exophiala ) dermatitidis is arguably the most studied phaeoid fungal pathogen of humans. Here, we report our comparative analysis of the genome of W. dermatitidis and the transcriptional response to low pH stress. This revealed that W. dermatitidis has lost the ability to synthesize alpha-glucan, a cell wall compound many pathogenic fungi use to evade the host immune system. In contrast, W. dermatitidis contains a similar profile of chitin synthase genes as related fungi and strongly induces genes involved in cell wall synthesis in response to pH stress. The large portfolio of transporters may provide W. dermatitidis with an enhanced ability to remove harmful products as well as to survive on diverse nutrient sources. The genome encodes three independent pathways for producing melanin, an ability linked to pathogenesis; these are active during pH stress, potentially to produce a barrier to accumulated oxidative damage that might occur under stress conditions. In addition, a full set of fungal light-sensing genes is present, including as part of a carotenoid biosynthesis gene cluster. Finally, we identify a two-gene cluster involved in nucleotide sugar metabolism conserved with a subset of fungi and characterize a horizontal transfer event of this cluster between fungi and algal viruses. This work reveals how W. dermatitidis has adapted to stress and survives in diverse environments, including during human infections.

Description: ELYS determines the subcellular localizations of Nucleoporins (Nups) during interphase and mitosis. We made loss-of-function mutations of Elys in Drosophila melanogaster and found that ELYS is dispensable for zygotic viability and male fertility but the maternal supply is necessary for embryonic development. Subsequent to fertilization, mitotic progression of the embryos produced by the mutant females is severely disrupted at the first cleavage division, accompanied by irregular behavior of mitotic centrosomes. The Nup160 introgression from D. simulans shows close resemblance to that of the Elys mutations, suggesting a common role for those proteins in the first cleavage division. Our genetic experiments indicated critical interactions between ELYS and three Nup107–160 subcomplex components; hemizygotes of either Nup37 , Nup96 or Nup160 were lethal in the genetic background of the Elys mutation. Not only Nup96 and Nup160 but also Nup37 of D. simulans behave as recessive hybrid incompatibility genes with D. melanogaster . An evolutionary analysis indicated positive natural selection in the ELYS-like domain of ELYS. Here we propose that genetic incompatibility between Elys and Nups may lead to reproductive isolation between D. melanogaster and D. simulans , although direct evidence is necessary.