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Novel and nodulation-regulated microRNAs in soybean roots.

Subramanian S, Fu Y, Sunkar R, Barbazuk WB, Zhu JK, Yu O - BMC Genomics (2008)

Bottom Line: Construction and analysis of a small RNA library led to the identification of 20 conserved and 35 novel miRNA families in soybean.The availability of complete and assembled genome sequence information will enable identification of many other miRNAs.The conserved miRNA loci and novel miRNAs identified in this study enable investigation of the role of miRNAs in rhizobial symbiosis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Donald Danforth Plant Science Center, 975 N Warson Road, St Louis, MO 63132, USA. ssubramanian@danforthcenter.org

ABSTRACT

Background: Small RNAs regulate a number of developmental processes in plants and animals. However, the role of small RNAs in legume-rhizobial symbiosis is largely unexplored. Symbiosis between legumes (e.g. soybean) and rhizobia bacteria (e.g. Bradyrhizobium japonicum) results in root nodules where the majority of biological nitrogen fixation occurs. We sought to identify microRNAs (miRNAs) regulated during soybean-B. japonicum symbiosis.

Results: We sequenced approximately 350000 small RNAs from soybean roots inoculated with B. japonicum and identified conserved miRNAs based on similarity to miRNAs known in other plant species and new miRNAs based on potential hairpin-forming precursors within soybean EST and shotgun genomic sequences. These bioinformatics analyses identified 55 families of miRNAs of which 35 were novel. A subset of these miRNAs were validated by Northern analysis and miRNAs differentially responding to B. japonicum inoculation were identified. We also identified putative target genes of the identified miRNAs and verified in vivo cleavage of a subset of these targets by 5'-RACE analysis. Using conserved miRNAs as internal control, we estimated that our analysis identified approximately 50% of miRNAs in soybean roots.

Conclusion: Construction and analysis of a small RNA library led to the identification of 20 conserved and 35 novel miRNA families in soybean. The availability of complete and assembled genome sequence information will enable identification of many other miRNAs. The conserved miRNA loci and novel miRNAs identified in this study enable investigation of the role of miRNAs in rhizobial symbiosis.

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Abundance of RNA sequences of different lengths in the library. After filtering for high-quality reads and trimming, the majority of the reads in the library ranged from 17 – 31 nt in length (white bars). The majority of the conserved miRNAs identified based on homology to sequences in miRbase ranged from 20–21 nt in length suggesting that our library was of high quality (see text).
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Figure 2: Abundance of RNA sequences of different lengths in the library. After filtering for high-quality reads and trimming, the majority of the reads in the library ranged from 17 – 31 nt in length (white bars). The majority of the conserved miRNAs identified based on homology to sequences in miRbase ranged from 20–21 nt in length suggesting that our library was of high quality (see text).

Mentions: A total of 3758 sequences in our library matched previously identified miRNAs listed in miRbase ('conserved miRNAs' in Figure 1B). About 68% of these reads were 20–24 nt in length representing possibly full length mature miRNAs (Figure 2). About 28% of the reads were 17–19 nt in length. These are very likely to be genuine miRNAs shortened due to cloning, trimming and/or sequencing artifacts. Majority of the conserved miRNAs in our libraries ranged from 20–21 nt in length (~60%; Figure 2) suggesting that we had high quality small RNA libraries.


Novel and nodulation-regulated microRNAs in soybean roots.

Subramanian S, Fu Y, Sunkar R, Barbazuk WB, Zhu JK, Yu O - BMC Genomics (2008)

Abundance of RNA sequences of different lengths in the library. After filtering for high-quality reads and trimming, the majority of the reads in the library ranged from 17 – 31 nt in length (white bars). The majority of the conserved miRNAs identified based on homology to sequences in miRbase ranged from 20–21 nt in length suggesting that our library was of high quality (see text).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2335117&req=5

Figure 2: Abundance of RNA sequences of different lengths in the library. After filtering for high-quality reads and trimming, the majority of the reads in the library ranged from 17 – 31 nt in length (white bars). The majority of the conserved miRNAs identified based on homology to sequences in miRbase ranged from 20–21 nt in length suggesting that our library was of high quality (see text).
Mentions: A total of 3758 sequences in our library matched previously identified miRNAs listed in miRbase ('conserved miRNAs' in Figure 1B). About 68% of these reads were 20–24 nt in length representing possibly full length mature miRNAs (Figure 2). About 28% of the reads were 17–19 nt in length. These are very likely to be genuine miRNAs shortened due to cloning, trimming and/or sequencing artifacts. Majority of the conserved miRNAs in our libraries ranged from 20–21 nt in length (~60%; Figure 2) suggesting that we had high quality small RNA libraries.

Bottom Line: Construction and analysis of a small RNA library led to the identification of 20 conserved and 35 novel miRNA families in soybean.The availability of complete and assembled genome sequence information will enable identification of many other miRNAs.The conserved miRNA loci and novel miRNAs identified in this study enable investigation of the role of miRNAs in rhizobial symbiosis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Donald Danforth Plant Science Center, 975 N Warson Road, St Louis, MO 63132, USA. ssubramanian@danforthcenter.org

ABSTRACT

Background: Small RNAs regulate a number of developmental processes in plants and animals. However, the role of small RNAs in legume-rhizobial symbiosis is largely unexplored. Symbiosis between legumes (e.g. soybean) and rhizobia bacteria (e.g. Bradyrhizobium japonicum) results in root nodules where the majority of biological nitrogen fixation occurs. We sought to identify microRNAs (miRNAs) regulated during soybean-B. japonicum symbiosis.

Results: We sequenced approximately 350000 small RNAs from soybean roots inoculated with B. japonicum and identified conserved miRNAs based on similarity to miRNAs known in other plant species and new miRNAs based on potential hairpin-forming precursors within soybean EST and shotgun genomic sequences. These bioinformatics analyses identified 55 families of miRNAs of which 35 were novel. A subset of these miRNAs were validated by Northern analysis and miRNAs differentially responding to B. japonicum inoculation were identified. We also identified putative target genes of the identified miRNAs and verified in vivo cleavage of a subset of these targets by 5'-RACE analysis. Using conserved miRNAs as internal control, we estimated that our analysis identified approximately 50% of miRNAs in soybean roots.

Conclusion: Construction and analysis of a small RNA library led to the identification of 20 conserved and 35 novel miRNA families in soybean. The availability of complete and assembled genome sequence information will enable identification of many other miRNAs. The conserved miRNA loci and novel miRNAs identified in this study enable investigation of the role of miRNAs in rhizobial symbiosis.

Show MeSH