Transcriptome Characteristics and Six Alternative Expressed Genes Positively Correlated with the Phase Transition of Annual Cambial Activities in Chinese Fir (<i>Cunninghamia lanceolata</i> (Lamb.) Hook)

<div><p>Background</p><p>The molecular mechanisms that govern cambial activity in angiosperms are well established, but little is known about these molecular mechanisms in gymnosperms. Chinese fir (<i>Cunninghamia lanceolata</i> (Lamb.) Hook), a diploid (2<i>n</i>  = 2<i>x</i>  = 22) gymnosperm, is one of the most important industrial and commercial timber species in China. Here, we performed transcriptome sequencing to identify the repertoire of genes expressed in cambium tissue of Chinese fir.</p><p>Methodology/Principal Findings</p><p>Based on previous studies, the four stage-specific cambial tissues of Chinese fir were defined using transmission electron microscopy (TEM). In total, 20 million sequencing reads (3.6 Gb) were obtained using Illumina sequencing from Chinese fir cambium tissue collected at active growth stage, with a mean length of 131 bp and a N50 of 90 bp. SOAPdenovo software was used to assemble 62,895 unigenes. These unigenes were further functionally annotated by comparing their sequences to public protein databases. Expression analysis revealed that the altered expression of six homologous genes (<i>ClWOX1</i>, <i>ClWOX4</i>, <i>ClCLV1</i>-<i>like</i>, <i>ClCLV</i>-<i>like</i>, <i>ClCLE12</i>, and <i>ClPIN1</i>-<i>like</i>) correlated positively with changes in cambial activities; moreover, these six genes might be directly involved in cambial function in Chinese fir. Further, the full-length cDNAs and DNAs for <i>ClWOX1</i> and <i>ClWOX4</i> were cloned and analyzed.</p><p>Conclusions</p><p>In this study, a large number of tissue/stage-specific unigene sequences were generated from the active growth stage of Chinese fir cambium. Transcriptome sequencing of Chinese fir not only provides extensive genetic resources for understanding the molecular mechanisms underlying cambial activities in Chinese fir, but also is expected to be an important foundation for future genetic studies of Chinese fir. This study indicates that <i>ClWOX1</i> and <i>ClWOX4</i> could be possible reverse genetic target genes for revealing the molecular mechanisms of cambial activities in Chinese fir.</p></div

Mapping of assembled unigenes of the Chinese fir transcriptome to KEGG pathways.

<p>Mapping of assembled unigenes of the Chinese fir transcriptome to KEGG pathways.</p

GO classification of the transcriptome.

<p>GO classification of the Chinese fir transcriptome. The horizontal axis indicates the three main categories that include molecular function, cellular component, and biological process, and certain categories that are self evident. The horizontal axis indicates the three main categories that include molecular function, cellular component, and biological process, and certain categories that are self evident. The right vertical axis indicates the number of assembled unigenes in each category, and the left vertical axis indicates the percentage of a certain type of subcategory in that main category.</p

Comparison of length distribution between hit and no-hit unigenes.

<p>The horizontal axis indicates length distribution of hit and no-hit assembled unigenes. The vertical axis indicates the percentage of hit and no-hit assembled unigenes.</p

Summary of the percentage of unigenes annotated for Chinese fir.

<p>Summary of the percentage of unigenes annotated for Chinese fir.</p

Random distribution of HiSeq 2000 sequencing reads in the assembled unigenes.

<p>The horizontal axis indicates relative position in gene (5'→3'). The vertical axis indicates the number of assembled reads.</p

COG functional classification of the transcriptome.

<p>A total of 15,662 unigenes were classified into 25 COG categories.</p

Expression analysis of 17 homologous genes in cambium tissues during the four growth stages, S1–S4, using qRT-PCR.

<p>Error bars represent the standard error of three biological replicates.</p

Length distribution of assembled contigs and unigenes.

<p>Length distribution of assembled contigs and unigenes.</p

The 17 homologous genes used for qRT-PCR.

<p>The 17 homologous genes used for qRT-PCR.</p