h6_2
Protocols: Mice were euthanized and colon tissues were harvested. The rectal tissues were discarded and the wound area identified with the help of a dissection microscope. The wound area was dissected out using a 3 millimeter standard biopsy puncher. Total RNA from the tissues were harvested using the column purification method using the Total RNA Kit, peqGOLD (VWR GmbH, Munich, Germnay) as per the manufacturer's instructions. From RNA samples extraction to final data obtained, every step of procedures containing samples testing, library construction and sequencing would directly affect the quality and quantity of data, which determines the bioinfomatic analysis results. Novogene promises to strictly control on all production data with high quality output and ensure the raw data accurate and reliable. Briefly, mRNA from Eukaryote organisms is purified from total RNA using poly-T oligo-attached magnetic beads (For prokaryotes, mRNA was purified through the removal ofrRNA by using kit). The mRNA is first fragmented randomly by addition of fragmentation buffer. Then first strand cDNA is synthesized using random hexamer primer and M-MuLV Reverse Transcriptase (RNase H-). Second strand cDNA synthesis is subsequently performed using DNA Polymerase I and RNase H. Double-stranded cDNA is purified using AMPure XP beads. Remaining overhangs of the purified double-stranded cDNA are converted into blunt ends via exonuclease/polymerase activities. After adenylation of 3' ends of DNA fragments, NEBNext Adaptor with hairpin loop structure is ligated to prepare for hybridization. In order to select cDNA fragments of preferentially 150~200 bp in length, the library fragments are purified with AMPure XP system (Beckma Coulter, Beverly, USA). Finally, the final library is gotten by PCR amplification and purification of PCR products by AMPure XP beads.
ENA first public
2021-11-05
ENA last update
2021-11-05
External Id
SAMEA9530996
INSDC center alias
Postdoctoral fellow
INSDC center name
Postdoctoral fellow
INSDC first public
2021-11-05T00:14:46Z
INSDC last update
2021-11-05T00:14:46Z
INSDC status
public
Submitter Id
E-MTAB-10824:h6_2
age
13
broker name
ArrayExpress
common name
house mouse
developmental stage
adult
genotype
wild type genotype
individual
5
organism part
descending colon
sample name
E-MTAB-10824:h6_2
sex
male
strain
C57/BL/6NRj