 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Sep 01, 2021 |
| Title |
small RNA KS1_2 |
| Sample type |
SRA |
| |
|
| Source name |
blood
|
| Organism |
Homo sapiens |
| Characteristics |
tumor origin: lung cancer (small cell carciinoma)
|
| Treatment protocol |
Total RNA was extracted from the tissues using Trizol (Invitrogen, Carlsbad, CA, USA), according to the manual instructions. For tissue samples, grind about 60mg with liquid nitrogen into powder and transfer the powder samples into the 2ml tube contains 1.5ml Trizol reagent. The mix was centrifuge at 12000×g for 5min at 4°C. The supernatant was transferred to a new 2.0ml tube which was added 0.3ml of Chloroform/isoamyl alcohol (24:1) per 1.5ml of Trizol reagent. After the mix was centrifuged at 12000×g for 10min at 4°C, the aqueous phase was transferred to a new 1.5mL tube which was add equal volume of supernatant of isopropyl alcohol. The mix was centrifuged at 12000×g for 20min at 4°C and then removed the supernatant. After washed with 1ml 75% ethanol, the RNA pellet was air-dried in the biosafety cabinet and then dissolved by add 25μL~100μL of DEPC-treated water. Subsequently, total RNA was qualified and quantified using a Nano Drop and Agilent 2100 bioanalyzer (Thermo Fisher Scientific, MA, USA)
|
| Growth protocol |
The samples (4ml blood) in PAXgene Blood RNA tube was handled and kept at –80 C until shipping to BGI Shenzhen by dry-ice cargo. The RNA extraction, quality check for sequencing were performed in BGI Shenzhen according to the BGI protocols.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Oligo(dT)-attached magnetic beads were used to purified mRNA. Purified mRNA was fragmented into small pieces with fragment buffer at appropriate temperature. Then First-strand cDNA was generated using random hexamer-primed reverse transcription, followed by a second-strand cDNA synthesis. afterwards, A-Tailing Mix and RNA Index Adapters were added by incubating to end repair. The cDNA fragments obtained from previous step were amplified by PCR, and products were purified by Ampure XP Beads, then dissolved in EB solution. The product was validated on the Agilent Technologies 2100 bioanalyzer for quality control. The double stranded PCR products from previous step were heated denatured and circularized by the splint oligo sequence to get the final library. The single strand circle DNA (ssCir DNA) was formatted as the final library. The final library was amplified with phi29 to make DNA nanoball (DNB) which had more than 300 copies of one molecular, DNBs were loaded into the patterned nanoarray and single end 50 bases reads were generated on BGIseq500 platform (BGI-Shenzhen, China).
Library was prepared with 1 μg total RNA for each sample. Total RNA was purified by electrophoretic separation on a 15% urea denaturing polyacrylamide gel electrophoresis (PAGE) gel and small RNA regions corresponding to the 18–30 nt bands in the marker lane (14-30 ssRNA Ladder Marker, TAKARA) were excised and recovered. Then the 18–30 nt small RNAs were ligated to a 5′-adaptor and a 3′-adaptor. The adapter-ligated small RNAs were subsequently transcribed into cDNA by SuperScript II Reverse Transcriptase (Invitrogen, USA) and then several rounds of PCR amplification with PCR Primer Cocktail and PCR Mix were performed to enrich the cDNA fragments. The PCR products were selected by agarose gel electrophoresis with target fragments 100~120 bp, and then purified by QIAquick Gel Extraction Kit (QIAGEN, Valencia, CA).
|
| |
|
| Library strategy |
ncRNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
BGISEQ-500 |
| |
|
| Description |
blood sample- 30 days after taking Huaier
stage IV, multiple metastasis
|
| Data processing |
RNA: Alignment:Hisat2 and Bowtie2
RNA: Expression of genes by RSEM
Call DEGs by PossionDis method
Predicted novel transcripts by StringTie and call SNP by GATK
hg38
Genome_build: TXT, FA
|
| |
|
| Submission date |
Aug 28, 2020 |
| Last update date |
Sep 01, 2021 |
| Contact name |
Manami Tanaka |
| E-mail(s) |
huaier_bradeioninstitute@hotmail.com
|
| Organization name |
Bradeion Institute of Medical Sciences
|
| Street address |
2-22-9 Hiratsuka, Kanagawa 259-1211, Japan
|
| City |
Kanagawa |
| ZIP/Postal code |
221-0045 |
| Country |
Japan |
| |
|
| Platform ID |
GPL23227 |
| Series (1) |
| GSE157086 |
Large-scale genomic alterations identified in cancer patients on the course of cancer recovery with complementary Huaier treatment identified by total RNA- and small non-coding RNA-sequencing. |
|
| Relations |
| BioSample |
SAMN15933778 |
| SRA |
SRX9031263 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4752561_KS1_2.miRNA.expr.txt.gz |
13.2 Kb |
(ftp)(http) |
TXT |
| GSM4752561_KS1_2_novelmiRNA.fa.gz |
2.6 Kb |
(ftp)(http) |
FA |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
|
|
|
|
 |
