Open Access Highly Accessed Research article

Strand-specific RNA-seq reveals widespread occurrence of novel cis-natural antisense transcripts in rice

Tingting Lu1*, Chuanrang Zhu1, Guojun Lu1, Yunli Guo1, Yan Zhou1, Zhiyong Zhang2, Yan Zhao1, Wenjun Li1, Ying Lu1, Weihua Tang2, Qi Feng1 and Bin Han1*

Author Affiliations

1 National Center for Gene Research & Institute of Plant Physiology and Ecology, Shanghai Institutes of Biological Sciences, Chinese Academy of Sciences, Shanghai, 200233, China

2 National Key Laboratory of Plant Molecular Genetics, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, 200233, China

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BMC Genomics 2012, 13:721  doi:10.1186/1471-2164-13-721

Published: 22 December 2012

Additional files

Additional file 1:

Summary of pair-end reads of ssRNA-seq and small RNAs from normal and three abiotic stress conditions.

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Additional file 2:

Statistics of rice transcripts.

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Additional file 3:

Summary of 5813 putative cis-NATs in rice. This table lists all cis-NATs identified from rice renewed assembled transcripts. Each column represents plus transcripts (transcriptional orientations are the same as the reference), its length, minus transcripts, length, overlapped length and detailed overlapped locations in rice genome (IRGSP v5.0) as indicated.

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Additional file 4:

Overview of small RNAs in rice. (A) Distribution of the lengths of all unique small RNAs generated by high-throughput sequencing from rice under four different conditions. (B) Length distribution of nat-siRNAs. (C) Distribution of all unique small RNAs located in different sequence components. (D) Distribution of nat-siRNAs located in different sequence components. The blue, red, green and purple bars represent small RNAs or nat-siRNAs from normal, salt stressed, cold stressed and drought stressed conditions, respectively (A-D). (E) First-nucleotide distribution of all unique small RNAs under four different conditions. (F) First-nucleotide distribution of nat-siRNAs under four different conditions.

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Additional file 5:

Sequences of small RNAs mapped to the overlapped regions ofcis-NATs. The 25,420, 18,598, 18,152 and 28,807 unique small RNAs from normal, salt, cold and drought conditions, respectively, are shown to be perfectly mapped to the overlapped regions of cis-NATs.

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Additional file 6:

Details of 2292 expressedcis-NATs. The detailed information of 2292 cis-NAT pairs with expression evidence (i.e. FPKM > 0) of both sense and antisense transcripts and with nat-siRNAs in the overlapping region under normal (1789 pairs), cold (1668), salt (1572) and drought (1668) conditions are shown. Columns 1-4 and 9-11 represent cis-NATs type, transcripts, protein domain and the length of transcripts, respectively. Columns 5-8 and 12-15 represent FPKM of transcripts under normal, salt, cold and drought conditions, respectively. Column 16 shows the overlapped length. Columns 17-20 show the number of corresponding nat-siRNAs located in the overlapped regions. The rest of the columns are the detailed strand bias of nat-siRNAs from four different conditions.

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Additional file 7:

Lists of 166cis-NATs that produced nat-siRNAs exclusively in the overlapping regions with more than five unique small RNAs from four different conditions. Of 166 cis-NATs, 72, 58, 66 and 75 cis-NATs were obtained from normal, salt, cold and drought treatments, respectively. These cis-NATs are listed separately in four tables, respectively indicated by blue, orange, red and green colour.

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Additional file 8:

Lists of 13cis-NATs that produced nat-siRNAs more enriched in the overlapping regions than the non-overlapping regions. Of 13 cis-NATs, nine, six, five and five cis-NATs obtained from normal, salt, cold and drought treatments, respectively. These cis-NATs are listed separately in four tables, respectively indicated by blue, orange, red and green colour.

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Additional file 9:

Strand bias ofcis-NATs that gave rise to nat-siRNAs. The strand bias is computed among different types of cis-NAT pairs under four different conditions. A total of nine types are listed. Here, plus indicates that the orientation of transcripts are the same as the reference genome, minus indicates opposite orientation of transcripts. One strand means only one strand of the cis-NATs gave rise to small RNAs. The plus/minus or minus/plus means one strand of the cis-NATs spawns at least five (part A) or two (part B) fold more small RNAs than the other.

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Additional file 10:

Scatter plots of expression of 1072cis-NATs. The two scatter plots compare transcripts expression ratio trends of 1072 co-expressed cis-NAT pairs between normal and drought-stress conditions (A), and between normal and salt-stress conditions (B). The results show five subgroups: red, green, purple, blue and orange spots represent subgroups 1-5, respectively.

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Additional file 11:

Lists of 112cis-NATs with DEGs. Differentially Expressed Genes (DEGs) were identified in one-to-one cis-NATs under salt, drought and cold treatments by comparing their FPKM with that under normal conditions. The gene names, FPKM, fold change (with absolute value > 2) and p-value (< 0.001) are listed.

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Additional file 12:

All many-to-manycis-NAT groups. In total, 223 cis-NAT groups were identified as many-to-many type. Of them, 209 groups belonged to one-to-two type, nine belonged to one-to-three type, four belonged to two-to-two groups, and one belonged to the one-to-four group. The meaning of each column is as given in Additional file 6.

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Additional file 13:

Statistics of 209 networks formed bycis-NAT groups. The numbers of one-to-two cis-NAT groups with expression evidence under normal, salt, cold and drought conditions are shown. These one-to-two cis-NAT groups were divided into five classes according to their cis-NATs types. Here, ‘All EXP’ indicates all cis-NAT groups with expression evidence (FPKM > 0) of both sense and antisense transcript, and with nat-siRNAs (number of small RNAs > 1) in overlapping region as well. ‘Either EXP’ indicates either of cis-NAT groups with expression evidence of both sense and antisense transcript, and with nat-siRNAs in the overlapping region as well.

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Additional file 14:

Primers designed for real-time RT-PCR and Northern Blots in this research. (DOCX 13 kb)

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