Transcriptional sequencing and analysis of major genes involved in the adventitious root formation of mango cotyledon segments

Main conclusion A total of 74,745 unigenes were generated and 1975 DEGs were identified. Candidate genes that may be involved in the adventitious root formation of mango cotyledon segment were revealed.Adventitious root formation is a crucial step in plant vegetative propagation, but the molecular mechanism of adventitious root formation remains unclear. Adventitious roots formed only at the proximal cut surface (PCS) of mango cotyledon segments, whereas no roots were formed on the opposite, distal cut surface (DCS). To identify the transcript abundance changes linked to adventitious root development, RNA was isolated from PCS and DCS at 0, 4 and 7 days after culture, respectively. Illumina sequencing of libraries generated from these samples yielded 62.36 Gb high-quality reads that were assembled into 74,745 unigenes with an average sequence length of 807 base pairs, and 33,252 of the assembled unigenes at least had homologs in one of the public databases. Comparative analysis of these transcriptome databases revealed that between the different time points at PCS there were 1966 differentially expressed genes (DEGs), while there were only 51 DEGs for the PCS vs. DCS when time-matched samples were compared. Of these DEGs, 1636 were assigned to gene ontology (GO) classes, the majority of that was involved in cellular processes, metabolic processes and single-organism processes. Candidate genes that may be involved in the adventitious root formation of mango cotyledon segment are predicted to encode polar auxin transport carriers, auxin-regulated proteins, cell wall remodeling enzymes and ethylene-related proteins. In order to validate RNA-sequencing results, we further analyzed the expression profiles of 20 genes by quantitative real-time PCR. This study expands the transcriptome information for Mangifera indica and identifies candidate genes involved in adventitious root formation in cotyledon segments of mango.

主要结论 共产生了74745个单基因，鉴定出1975个差异表达基因。揭示了可能参与芒果子叶切段不定根形成的候选基因。不定根形成是植物营养繁殖的关键步骤，但不定根形成的分子机制仍不清楚。不定根仅在芒果子叶切段的近端切割面（PCS）形成，而在相对的远端切割面（DCS）没有根形成。为了确定与不定根发育相关的转录本丰度变化，分别在培养0、4和7天后从PCS和DCS中分离RNA。对这些样本构建的文库进行Illumina测序，产生了62.36 Gb的高质量读数，这些读数组装成74745个单基因，平均序列长度为807个碱基对，并且33252个组装的单基因至少在一个公共数据库中有同源物。对这些转录组数据库的比较分析表明，在PCS的不同时间点之间有1966个差异表达基因（DEGs），而在时间匹配的样本比较时，PCS与DCS之间只有51个DEGs。在这些DEGs中，1636个被归类到基因本体（GO）类别，其中大多数涉及细胞过程、代谢过程和单细胞生物过程。可能参与芒果子叶切段不定根形成的候选基因预计编码极性生长素运输载体、生长素调节蛋白、细胞壁重塑酶和乙烯相关蛋白。为了验证RNA测序结果，我们通过实时定量PCR进一步分析了20个基因的表达谱。这项研究扩展了芒果（Mangifera indica）的转录组信息，并确定了参与芒果子叶切段不定根形成的候选基因。