SNP Identification from RNA Sequencing and Linkage Map Construction of Rubber Tree for Anchoring the Draft Genome

<div><p><i>Hevea brasiliensis</i>, or rubber tree, is an important crop species that accounts for the majority of natural latex production. The rubber tree nuclear genome consists of 18 chromosomes and is roughly 2.15 Gb. The current rubber tree reference genome assembly consists of 1,150,326 scaffolds ranging from 200 to 531,465 bp and totalling 1.1 Gb. Only 143 scaffolds, totalling 7.6 Mb, have been placed into linkage groups. We have performed RNA-seq on 6 varieties of rubber tree to identify SNPs and InDels and used this information to perform target sequence enrichment and high throughput sequencing to genotype a set of SNPs in 149 rubber tree offspring from a cross between RRIM 600 and RRII 105 rubber tree varieties. We used this information to generate a linkage map allowing for the anchoring of 24,424 contigs from 3,009 scaffolds, totalling 115 Mb or 10.4% of the published sequence, into 18 linkage groups. Each linkage group contains between 319 and 1367 SNPs, or 60 to 194 non-redundant marker positions, and ranges from 156 to 336 cM in length. This linkage map includes 20,143 of the 69,300 predicted genes from rubber tree and will be useful for mapping studies and improving the reference genome assembly.</p></div

Linkage groups, the number of SNPs they contain, the number of non-redundant marker positions and the total length in cM.

<p>Linkage groups, the number of SNPs they contain, the number of non-redundant marker positions and the total length in cM.</p

Rubber tree linkage group ideograms with positions of non-redundant markers indicated as orange vertical bars and markers from within placed scaffolds indicated as blue vertical bars

<p>Rubber tree linkage group ideograms with positions of non-redundant markers indicated as orange vertical bars and markers from within placed scaffolds indicated as blue vertical bars</p

High-density genetic map of cassava.

<p>The SNP-based map included 2,110 non-redundant markers distributed over 19 linkage groups (LGs). The length of the each linkage group (in cM) was indicated at the bottom.</p

Analysis of synonymous and non-synonymous changes in cassava SNPs.

<p>Detailed information regarding the changes introduced by nucleotide substitutions and their distribution according to the nucleotide position within codons.</p><p>Analysis of synonymous and non-synonymous changes in cassava SNPs.</p

Manhattan plot of QTL LOD scores for starch viscosity traits.

<p>The Manhattan plots display the LOD scores of SNP markers along each linkage group (designated at the bottom). The red and orange lines indicated genome-wide and chromosome-wide significance thresholds, respectively.</p

Large-Scale SNP Discovery through RNA Sequencing and SNP Genotyping by Targeted Enrichment Sequencing in Cassava (<i>Manihot esculenta</i> Crantz)

<div><p>Cassava (<i>Manihot esculenta</i> Crantz) is one of the most important crop species being the main source of dietary energy in several countries. Marker-assisted selection has become an essential tool in plant breeding. Single nucleotide polymorphism (SNP) discovery via transcriptome sequencing is an attractive strategy for genome complexity reduction in organisms with large genomes. We sequenced the transcriptome of 16 cassava accessions using the Illumina HiSeq platform and identified 675,559 EST-derived SNP markers. A subset of those markers was subsequently genotyped by capture-based targeted enrichment sequencing in 100 F₁ progeny segregating for starch viscosity phenotypes. A total of 2,110 non-redundant SNP markers were used to construct a genetic map. This map encompasses 1,785 cM and consists of 19 linkage groups. A major quantitative trait locus (QTL) controlling starch pasting properties was identified and shown to coincide with the QTL previously reported for this trait. With a high-density SNP-based linkage map presented here, we also uncovered a novel QTL associated with starch pasting time on LG 10.</p></div

Distribution of SNP markers on the linkage groups.

<p>Distribution of SNP markers on the linkage groups.</p

Statistics for RNA-seq data from 16 cassava accessions.

<p>Quality scores measure the probability that the base is called incorrectly. Quality scores of 20 (Q20) represent a corresponding call accuracy of 99%.</p><p>Statistics for RNA-seq data from 16 cassava accessions.</p