|
| Status |
Public on Jun 23, 2020 |
| Title |
mouse_hippocampus_18mo_FAdeficient_Replicate1 |
| Sample type |
RNA |
| |
|
| Source name |
mouse hippocampus, 18 months folic acid deficiency
|
| Organism |
Mus musculus |
| Characteristics |
strain: CD1
age: 18 months
gender: female
tissue: hippocampus
sample type: folic acid deficiency
|
| Treatment protocol |
FA mice were give AIN93G normal folic acid (2 mg/ml), and DEF mice were deprived of all folate beginning post-weaning
|
| Growth protocol |
mice were kept on a 12 hr light/dark cycle in a temperature controlled room for the duration of the study.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
hippocampi were soaked in RNALater for 24 hr at 4 Celcius before being stored at -80 Celcius until ready for extraction. RNA was extracted using Trizol (Invitrogen) with the standard protocol provided by the manufacturer
|
| Label |
Cy3
|
| Label protocol |
Sample labeling and array hybridization were performed according to the Agilent One-Color Microarray-Based Gene Expression Analysis protocol (Agilent Technology) with minor modifications. Briefly, mRNA was purified from total RNA after removal of rRNA (mRNA-ONLY™ Eukaryotic mRNA Isolation Kit, Epicentre). Then, each sample was amplified and transcribed into fluorescent cRNA along the entire length of the transcripts without 3’ bias utilizing a mixture of oligo(dT) and random primers (Arraystar Flash RNA Labeling Kit, Arraystar). The labeled cRNAs were purified by RNeasy Mini Kit (Qiagen). The concentration and specific activity of the labeled cRNAs (pmol Cy3/μg cRNA) were measured by NanoDrop ND-1000.
|
| |
|
| Hybridization protocol |
1 μg of each labeled cRNA was fragmented by adding 5 μl 10 × Blocking Agent and 1 μl of 25 × Fragmentation Buffer, then heated the mixture at 60 °C for 30 min, finally 25 μl 2 × GE Hybridization buffer was added to dilute the labeled cRNA. 50 μl of hybridization solution was dispensed into the gasket slide and assembled to the LncRNA expression microarray slide. The slides were incubated for 17 hours at 65°C in an Agilent Hybridization Oven. The hybridized arrays were washed, fixed and scanned with using the Agilent DNA Microarray Scanner (part number G2505C).
|
| Scan protocol |
Agilent DNA Microarray Scanner (part number G2505C).
|
| Description |
Gene expression after 18 months of folic acid deficiency
|
| Data processing |
Agilent Feature Extraction software (version 11.0.1.1) was used to analyze acquired array images. Quantile normalization and subsequent data processing were performed with using the GeneSpring GX v12.1 software package (Agilent Technologies). After quantile normalization of the raw data, LncRNAs and mRNAs that at least 6 out of 12 samples have flags in Present or Marginal (“All Targets Value”) were chosen for further data analysis. Differentially expressed LncRNAs and mRNAs with statistical significance were identified through Volcano Plot filtering between the two groups of samples. Hierarchical Clustering was performed using the R software (version 2.15). GO analysis and Pathway analysis were performed in the standard enrichment computation method
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| |
|
| Submission date |
Apr 06, 2020 |
| Last update date |
Jun 23, 2020 |
| Contact name |
Abigail Lawton |
| E-mail(s) |
alenz@liberty.edu
|
| Organization name |
Liberty University
|
| Department |
Biology and Chemistry
|
| Lab |
Gary Isaacs
|
| Street address |
1971 University Blvd
|
| City |
Lynchburg |
| State/province |
Virginai |
| ZIP/Postal code |
24515 |
| Country |
USA |
| |
|
| Platform ID |
GPL25015 |
| Series (1) |
| GSE148126 |
Chronic Folate Deficiency in the Mouse Hippocampus |
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