Full-length cDNA libraries (< 1 kb, 1–2 kb, 2–3 kb, 3–6 kb size ranges) were generated from latex of a Hevea brasiliensis tree clone, RRIM 600, for PacBio Iso-Seq sequencing. The total number of high-quality consensus sequences obtained from nearly 20 Gb of raw reads was 207,943 having an average length of 1823 bp and N50 of 2538 bp. Mapping of high-quality consensus sequences to the published Reyan 7-33-97 genome sequence produced 37,224 Iso-Seq transcripts with an average length of 2030 bp and N50 of 2529 bp. These represented 14,999 genic loci. Comparison with the Reyan 7-33-97 gene models indicated that 50.75% of Iso-Seq transcripts were potentially novel gene isoforms. Subsequently, we also demonstrate the application of Iso-Seq in identifying previously unknown alternatively spliced transcripts based on three case studies of candidate rubber genes of interest. Findings from this study support the usefulness of PacBio Iso-Seq as transcriptome resources for gene family characterisation and genome annotation in Hevea.

中文翻译：

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PacBio Iso-Seq在橡胶树胶乳中进行转录本和基因发现的实用程序

从巴西橡胶树的乳胶中产生全长cDNA文库（<1 kb，1-2 kb，2-3 kb，3-6 kb）。树克隆RRIM 600，用于PacBio Iso-Seq测序。从近20 Gb的原始读取中获得的高质量共有序列的总数为207,943，平均长度为1823 bp，N50为2538 bp。将高质量共有序列映射到已发布的Reyan 7-33-97基因组序列可产生37,224个Iso-Seq转录本，平均长度为2030 bp，N50为2529 bp。这些代表了14,999个基因位点。与Reyan 7-33-97基因模型的比较表明，50.75％的Iso-Seq转录本是潜在的新型基因亚型。随后，我们还基于候选橡胶基因的三个案例研究，证明了Iso-Seq在鉴定以前未知的可变剪接转录本中的应用。橡胶树。