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| Status |
Public on Nov 01, 2014 |
| Title |
MiRNAs in tobacco roots after topping |
| Sample type |
SRA |
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| Source name |
tobacco roots after topping
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| Organism |
Nicotiana tabacum |
| Characteristics |
cultivar: K326
tissue: roots
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| Treatment protocol |
Topping treatment
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| Growth protocol |
Tobacco plants were grown in the fields.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated from roots tissue using Trizol agent (TaKaRa, Dalian, China), according to the manufacturer’s instructions. MiRNA cloning was performed as described previously by Sunkar and Zhu [23]. Briefly, 0.5 M NaCl and 10% PEG8000 were used to precipitate and enrich RNAs with low molecular weight. Next, a 15% polyacrylamide denaturing gel was employed to separate the low-molecular weight RNA. During gel electrophoresis, about 100 μg of total RNA was applied to the gel and two labeled RNA oligonucleotides, approximately 18 and 26 nt in length, were used as size standards. After gel electrophoresis, small RNAs with 18-26 nt were excised from the gel and eluted with 0.4 M NaCl overnight at 4°C. The RNA was dephosphorylated using alkaline phosphatase (New England Biolabs, Beijing China) and recovered by ethanol precipitation. The isolated small RNAs were then sequentially ligated to 5’- and 3’-chimeric oligonucleotide adapters, reversely transcribed, and amplified by PCR. Finally, Solexa sequencing technology was employed to sequence the small RNAs from tobacco roots samples (Beijing Genomics Institute, Shenzhen, Guangdong, China).
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina HiSeq 2000 |
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| Data processing |
Only small RNA reads that passed the Illumina pipeline quality control and contained clear adaptor sequences were considered good reads for further processing. After adaptor sequence was trimmed, clean small RNA reads of 18nt or more were combined into unique sequences. Reads that match known plant repeats, rRNAs, tRNAs, snRNAs, and snoRNAs were removed. The unique small RNA reads were used to do a Blastn search against the four genomic sequence resources：317, 49 tobacco ESTs (www.ncbi.nlm.nih.gov)，300,158 tobacco genomic sequences (ftp.solgenomics.net/tobacco_genome), 44,561 tobacco shotgun sequences (ftp.tigr.org/pub/data/plantta/Nicotiana_tabacum), and 1,420,595 tobacco Genome Survey Sequences (GSS，www.ncbi.nlm.nih.gov). Perfect match was required. Results of these BLASTn searches were then subjected to secondary structure analysis using UNAFOLD version 3.8. We then examined the secondary structure, and a result was considered as a genuine miRNA candidate if it met the miRNA criteria [24]. To identify the conserved miRNAs in tobacco, these miRNA candidates were used to do a Blastn search against the miRNA database (miRBase17.0, http://www.mirbase.org), and only the perfectly matched sequences (number of mismatch<3) were considered to be conserved miRNAs. Except for these conserved miRNAs, other miRNA candidates were considered to be new miRNAs. The sequence reads of each miRNA in two libraries were normalized, and then the fold change of miRNAs between two libraries was obtained.
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| Submission date |
Nov 01, 2011 |
| Last update date |
May 15, 2019 |
| Contact name |
hongxiang guo |
| E-mail(s) |
guohongxiang06@126.com
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| Organization name |
henan agricultural university
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| Street address |
95 wenhua road
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| City |
zhengzhou |
| ZIP/Postal code |
450002 |
| Country |
China |
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| Platform ID |
GPL13760 |
| Series (1) |
| GSE33370 |
Differential expression of miRNAs in response to topping in tobacco roots |
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| Relations |
| SRA |
SRX104051 |
| BioSample |
SAMN00749817 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM825366_clean_after_topping.fa.gz |
30.1 Mb |
(ftp)(http) |
FA |
| GSM825366_clean_after_topping.txt.gz |
24.1 Mb |
(ftp)(http) |
TXT |
| GSM825366_lengthVSnumber_after_topping.txt.gz |
199 b |
(ftp)(http) |
TXT |
| GSM825366_small_RNA_in_roots_after_topping.txt.gz |
2.9 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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