Era of gapless plant genomes: innovations in sequencing and mapping technologies revolutionize genomics and breeding

Whole-genome sequencing and assembly have revolutionized plant genetics and molecular biology over the last two decades. However, significant shortcomings in first- and second-generation technology resulted in imperfect reference genomes: numerous and large gaps of low quality or undeterminable sequence in areas of highly repetitive DNA along with limited chromosomal phasing restricted the ability of researchers to characterize regulatory noncoding elements and genic regions that underwent recent duplication events. Recently, advances in long-read sequencing have resulted in the first gapless, telomere-to-telomere (T2T) assemblies of plant genomes. This leap forward has the potential to increase the speed and confidence of genomics and molecular experimentation while reducing costs for the research community

Ten new high-quality genome assemblies for diverse bioenergy sorghum genotypes

INTRODUCTION: Sorghum (Sorghum bicolor (L.) Moench) is an agriculturally and economically important staple crop that has immense potential as a bioenergy feedstock due to its relatively high productivity on marginal lands. To capitalize on and further improve sorghum as a potential source of sustainable biofuel, it is essential to understand the genomic mechanisms underlying complex traits related to yield, composition, and environmental adaptations. METHODS: Expanding on a recently developed mapping population, we generated de novo genome assemblies for 10 parental genotypes from this population and identified a comprehensive set of over 24 thousand large structural variants (SVs) and over 10.5 million single nucleotide polymorphisms (SNPs). RESULTS: We show that SVs and nonsynonymous SNPs are enriched in different gene categories, emphasizing the need for long read sequencing in crop species to identify novel variation. Furthermore, we highlight SVs and SNPs occurring in genes and pathways with known associations to critical bioenergy-related phenotypes and characterize the landscape of genetic differences between sweet and cellulosic genotypes. DISCUSSION: These resources can be integrated into both ongoing and future mapping and trait discovery for sorghum and its myriad uses including food, feed, bioenergy, and increasingly as a carbon dioxide removal mechanism

Gene disruption by structural mutations drives selection in US rice breeding over the last century.

The genetic basis of general plant vigor is of major interest to food producers, yet the trait is recalcitrant to genetic mapping because of the number of loci involved, their small effects, and linkage. Observations of heterosis in many crops suggests that recessive, malfunctioning versions of genes are a major cause of poor performance, yet we have little information on the mutational spectrum underlying these disruptions. To address this question, we generated a long-read assembly of a tropical japonica rice (Oryza sativa) variety, Carolina Gold, which allowed us to identify structural mutations (>50 bp) and orient them with respect to their ancestral state using the outgroup, Oryza glaberrima. Supporting prior work, we find substantial genome expansion in the sativa branch. While transposable elements (TEs) account for the largest share of size variation, the majority of events are not directly TE-mediated. Tandem duplications are the most common source of insertions and are highly enriched among 50-200bp mutations. To explore the relative impact of various mutational classes on crop fitness, we then track these structural events over the last century of US rice improvement using 101 resequenced varieties. Within this material, a pattern of temporary hybridization between medium and long-grain varieties was followed by recent divergence. During this long-term selection, structural mutations that impact gene exons have been removed at a greater rate than intronic indels and single-nucleotide mutations. These results support the use of ab initio estimates of mutational burden, based on structural data, as an orthogonal predictor in genomic selection

Low-phosphate-selected Auxenochlorella protothecoides redirects phosphate to essential pathways while producing more biomass

Despite the capacity to accumulate similar to 70% w/w of lipids, commercially produced unicellular green alga A. protothecoides may become compromised due to the high cost of phosphate fertilizers. To address this limitation A. protothecoides was selected for adaptation to conditions of 100x and 5x lower phosphate and peptone, respectively, compared to wild-type media'. The A. protothecoides showed initial signs of adaptation by 45-50days, and steady state growth at similar to 100 days. The low phosphate (P)-adapted strain produced up to similar to 30% greater biomass, while total lipids (similar to 10% w/w) remained about the same, compared to the wild-type strain. Metabolomic analyses indicated that the low P -adapted produced 3.3-fold more saturated palmitic acid (16:0) and 2.2-fold less linolenic acid (18:3), compared to the wild-type strain, resulting in an similar to 11% increase in caloric value, from 19.5kJ/g for the wild type strain to 21.6kJ/g for the low P -adapted strain, due to the amounts and composition of certain saturated fatty acids, compared to the wild type strain. Biochemical changes in A. protothecoides adapted to lower phosphate conditions were assessed by comparative RNA-Seq analysis, which yielded 27,279 transcripts. Among them, 2,667 and 15 genes were significantly down- and up-regulated, at >999-fold and >3-fold (adjusted p-value <0.1), respectively. The expression of genes encoding proteins involved in cellular processes such as division, growth, and membrane biosynthesis, showed a trend toward down-regulation. At the genomic level, synonymous SNPs and Indels were observed primarily in coding regions, with the 40S ribosomal subunit gene harboring substantial SNPs. Overall, the adapted strain out-performed the wild-type strain by prioritizing the use of its limited phosphate supply for essential biological processes. The low P-adapted A. protothecoides is expected to be more economical to grow over the wild-type strain, based on overall greater productivity and caloric content, while importantly, also requiring 100-fold less phosphate.National Alliance for Advanced Biofuels and Bioproducts (NAABB)Open access journal.This item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

Genotyping by sequencing of rice interspecific backcross inbred lines identifies QTLs for grain weight and grain length

Grain weight and grain length are the most stable components of rice yield and important indicators of consumer preference. Considering the potentials of wild rice and to enhance the rice yields to meet the increasing demands, 185 Backcross Inbred Lines (BILs) in the background of O. sativa ssp. indica cv. PR114, including 63 rufi-BILs derived from O. rufipogon IRGC104433 and 122 glumae-BILs from O. glumaepatula IRGC104387 were evaluated for mapping QTLs for yield and yield component traits using Genotyping by Sequencing (GBS). Phenotypic evaluation of BILs in three seasons spanning two locations revealed significant differences compared with recurrent parent. BILs which did not show significant differences for any trait under investigation, or similar based on pedigree, were excluded from GBS. Some glumae-BILs had to be excluded from mapping QTLs due to less sequence information. A custom designed approach for GBS data analysis identified 3322 informative SNPs in 55 rufi-BILs and 3437 informative SNPs in 79 glumae-BILs. QTL mapping identified one QTL for thousand grain weight (qtgw5.1), two for grain width (qgw5.1, qgw5.2) and one for grain length (qgl7.1) in rufi-BILs. In the glumae-BILs, three QTL for thousand grain weight (qtgw2.1, qtgw3.1, qtgw6.1) and two for grain length (qgl3.1, qgl7.1) were identified. Most of the grain weight and width QTL showed positive additive effect contributed by wild species allele, whereas the grain length QTL showed positive additive effect contributed by recurrent parent allele. Based on their physical position, none of the QTLs were found similar to previously cloned QTLs. QTLs for grain traits identified from low yielding wild relatives of rice reveals their significance in improving further the rice yields and widen the genetic base of cultivated rice

The fatty acid elongation and degradation KEGG pathway showing the role of dehydrogenase (E.C. 1.1.1.35; 5374-fold decrease) in fatty acid elongation and degradation.

<p>The fatty acid elongation and degradation KEGG pathway showing the role of dehydrogenase (E.C. 1.1.1.35; 5374-fold decrease) in fatty acid elongation and degradation.</p

Low-phosphate-selected <i>Auxenochlorella protothecoides</i> redirects phosphate to essential pathways while producing more biomass - Fig 6

<p><b>A</b>. A cluster heat map displaying the change of gene expression. Each column represented a different selection stage and the each low represented the level of transcription expressed in the color scheme. In total, 27,279 transcripts were compared across the selection process. The transcript levels observed at wk 2 were considered as the basal transcriptional level for comparison with gene expression levels observed at 6, 10, >14, and >22 wks. In general, transcripts were hierachically clustered, based on analysis using the Centroid method and Euclidean distance metric. <b>B.</b> The line graphs shows different gene expression patterns among the 27,279 transcripts, indicative of low phosphate selection. Among the 27,279 transcripts, >99.9% transcripts showed a gradual decrease in gene expression (line graph #1,#2, and #4), whereas, increased expression was observed for only <0.1% of transcripts (line graph #7 and #8).</p

Selected transcripts significantly suppressed in the low P-adapted <i>A</i>. <i>protothecoides</i> strain at >22 wks.

<p>Selected transcripts significantly suppressed in the low P-adapted <i>A</i>. <i>protothecoides</i> strain at >22 wks.</p

Lipid composition of wild-type and low P-adapted strain by LC-MS analysis.

<p><b>A</b>. Fatty acyls and saccharolipids were produced at similar levels by both strains (t-test, n = 3, P>0.05). <b>B</b>. Glycerophospholipid and polyketide accumulation was less in the low P-adapted compared to wild type strain (t-test, n = 3, P<0.05). <b>C</b>. Prenol lipids, glycerolipids, sphingolipids, and sterol lipids were increased in the low P-adapted strain (t-test, n = 3, P<0.05).</p

Flow cytometric analysis of wild-type and low P-adapted <i>A</i>. <i>protothecoides</i>.

<p>Density plots and corresponding histograms of wild-type (<b>A</b>) and low P-adapted (<b>B</b>) were shown for the chlorophyll-based fluorescence. <b>C.</b> Density plot and histogram of flow cytometric measurement were based on BODIPY fluorescence.</p