View Project rosen-illu-mouse-588443

Project Summary
Status:	Public
Publications:	1 Published
Project Detail		Data Detail
Platform:	Illumina	MIAME Areas	Compliance
Species:	Mouse	Array Design Detail	false
Organ/Tissue Type:	cerebral cortex	Experiment Detail	true
Organ Region:		Sample Detail	true
Cell Type:	brain tissue	Hybridization Detail	false
Study Type:	time_series_design	Measurement Detail	true
Disease/Condition:	normal
Replicates:	2
Expected Samples:	48

Available Actions

Investigator Contact Detail
Name	Glenn D Rosen
Institution:	Beth Israel Deaconess Medical Center
Street Address:	Department of Neurology
	330 Brookline Ave.,
City, State/Province:	Boston , MA
Zip/Postal Code:	02215
Country:	United States
Work Phone:	617-667-3252
Fax:	617-667-5217
E-mail:	grosen@bidmc.harvard.edu

Proposal Detail

Grant:

NS052397

Status:

Public

Service Type:

Hybrization through Analysis

IACUC:

036-2006

IACUC date:

2007-05-07

Study Relevance:

The neocortex is among the most studied structures in the brain, which is not surprising given its importance to cognitive and motor functioning. Changes in the organization of the neocortex, especially during development, can have a profound impact on cognitive capacity. Understanding the genes that modulate the development of this structures would have a strong impact on our understanding of a variety of developmental disorders, including autism and developmental dyslexia. Although neurogenesis in the neocortex is well studied, there is surprisingly little known about the processes that regulate the variation in the number of neurons and glia among individuals. Similarly, there is little understanding of the factors that modulate the variation in the volume of this structure. The studies proposed here will provide the foundation for these investigations.

Hypothesis:

We hypothesize that separate and distinct QTLs modulate variation in regional volume as opposed to the number of neurons and glia in the neocortex.

Specific Aim:

We aim to o map QTLs that selectively modulate glial and neuronal populations in the neocortex. The parent strains of this BXD recombinant inbred (RI) set (C57BL/6J and DBA/2J) have been genotyped, and genome-wide haplotype maps are available in GeneNetwork <www.genenetwork.org> to enhance identification of candidate genes within QTL intervals. Here will will take the first steps toward the creation of a publicly available database of developmental neocortical RNA transcript levels in GeneNetwork that will further boost our efforts to select candidate genes.

Experimental Procedure and Design:

We have dissected the neocortex from both males and females of parent strains of the BXD RI (C57BL/6J and DBA/2J) at postnatal ages (P)1, P3, P5, P7, P10, and P14. There are, therefore, 24 bio-sources in this experiment (2 strains X 2 sexes X 6 ages). At each age, a minimum of three litters were taken and tissue for RNA extraction was pooled among these three litters. There are two biological replicates for each bio-source for a total of 48 samples. Subjects were sacrificed by decapitation, and their brains quickly removed, bisected mid-sagittally, and placed in RNAlater for 15–20 minutes. This allowed the brain to harden slightly and eased the process of regional identification and dissection. The hemispheres are removed individually from the RNAlater, and the cerebellum, olfactory bulbs, and hippocampus are dissected and stored in RNAlater for the use of collaborators. The striatum and thalamic tissue was dissected away and what was left is the cerebral cortex (along with some white matter). After the completion of each dissection, the tissue was placed immediately into a fresh vial of RNAlater and stored at -20°C until the RNA is extracted. Dissected tissues were processed to extract total RNA using standard phenol-based protocols. Briefly, tissue was homogenized in Trizol. RNA was extracted with 0.2 ml chloroform/ml Trizol and centrifuged at 12,000g for 15 minutes. The RNA is precipitated from the aqueous phase in 0.5 ml isopropanol/ml and centrifuged for 10 minutes at 12,000g. The supernatant is removed, and the RNA pellet is washed in 75% ethanol, dried, and dissolved in DEPCtreated water.

Quality Control Description:

The quality and purity of RNA is assessed using spectrophotometry and gel electrophoresis. All RNA in this experiment had A260/280 ratios of 1.8 or above. There were technical replicates for each sample undergoing spectrophotometry.

Quality Control Types:

technical_replicate

Replicate Description:

A minimum of three litters were sacrificed at each age, and each litter had 4-9 pups and tissue was segregated by sex. To create a sample, tissue was pooled from each of the litters representing the strain, sex, and age of interest. A separate sample was created from the same litters as a biological replicate.

Replicate Types:

biological_replicate

Experimental Factors:

Conditions that are tested in the experiment. At least one is required. Experimental factors are the independent variables in the experiment.

Factor Name	Description	Factor Category
Age	P1, P3, P5, P7, P10, P14	age
Sex	Male, Female	sex
Strain	C57BL/6J, DBA/2J	strain_or_line

Project Samples

Samples associated with this project.

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Name	Description	Bio-Source	Extracts
RNA87	D2-P7 Male Sample 1	D2 - P7 Male	1
RNA88	D2-P7 Male Sample 2	D2 - P7 Male	1
RNA89	D2-P3 Male Sample 1	D2 - P3 Male	1
RNA49	D2-P1 Male Sample1	D2 - P1 Male	1
RNA50	D2-P1 Male Sample2	D2 - P1 Male	1
RNA51	B6-P7 Female Sample 1	B6 - P7 Female	1
RNA52	B6-P7 Female Sample 2	B6 - P7 Female	1
RNA53	B6-P1 Male Sample 1	B6 - P1 Male	1
RNA54	B6-P1 Male Sample 2	B6 - P1 Male	1
RNA55	B6-P3 Male Sample 1	B6 - P3 Male	1
RNA56	B6-P3 Male Sample 2	B6 - P3 Male	1
RNA57	B6-P5 Male Sample 1	B6 - P5 Male	1
RNA58	B6-P5 Male Sample 2	B6 - P5 Male	1
RNA59	B6-P7 Male Sample 1	B6 - P7 Male	1
RNA60	B6-P7 Male Sample 2	B6 - P7 Male	1
RNA61	B6-P10 Male Sample 1	B6 - P10 Male	1
RNA62	B6-P10 Male Sample 2	B6 - P10 Male	1
RNA63	B6-P1 Female Sample 1	B6 - P1 Female	1
RNA64	B6-P1 Female Sample 2	B6 - P1 Female	1
RNA65	B6-P3 Female Sample 1	B6 - P3 Female	1
RNA66	B6-P3 Female Sample 2	B6 - P3 Female	1
RNA67	B6-P5 Female Sample 1	B6 - P5 Female	1
RNA68	B6-P5 Female Sample 2	B6 - P5 Female	1
RNA69	B6-P10 Female Sample 1	B6 - P10 Female	1
RNA70	B6-P10 Female Sample 2	B6 - P10 Female	1
RNA71	B6-P14 Male Sample 1	B6- P14 Male	1
RNA72	B6-P14 Male Sample 2	B6- P14 Male	1
RNA73	B6-P14 Female Sample 1	B6 - P14 Female	1
RNA74	B6-P14 Female Sample 2	B6 - P14 Female	1
RNA75	D2-P1 Female Sample 1	D2 - P1 Female	1
RNA76	D2-P1 Female Sample 2	D2 - P1 Female	1
RNA77	D2-P5 Female Sample 1	D2 - P5 Female	1
RNA78	D2-P5 Female Sample 2	D2 - P5 Female	1
RNA79	D2-P7 Female Sample 1	D2 - P7 Female	1
RNA80	D2-P7 Female Sample 2	D2 - P7 Female	1
RNA81	D2-P10 Female Sample 1	D2 - P10 Female	1
RNA82	D2-P10 Female Sample 2	D2 - P10 Female	1
RNA83	D2-P14 Female Sample 1	D2 - P14 Female	1
RNA84	D2-P14 Female Sample 2	D2 - P14 Female	1
RNA85	D2-P5 Male Sample 1	D2 - P5 Male	1
RNA86	D2-P5 Male Sample 2	D2 - P5 Male	1
RNA90	D2-P3 Male Sample 2	D2 - P3 Male	1
RNA91	D2-P3 Female Sample 1	D2 - P3 Female	1
RNA92	D2-P3 Female Sample 2	D2 - P3 Female	1
RNA93	D2-P10 Male Sample 1	D2 - P10 Male	1
RNA94	D2-P10 Male Sample 2	D2 - P10 Male	1
RNA95	D2-P14 Male Sample 1	D2 - P14 Male	1
RNA96	D2-P14 Male Sample 2	D2 - P14 Male	1

Project Hybridizations

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Name	Labeled Extract	Hybridization Protocol
Hybridization48	RNA90_le1	Illumina
Hybridization49	RNA58_le1	Illumina
Hybridization50	RNA60_le1	Illumina
Hybridization51	RNA79_le1	Illumina
Hybridization52	RNA81_le1	Illumina
Hybridization53	RNA73_le1	Illumina
Hybridization54	RNA82_le1	Illumina
Hybridization55	RNA53_le1	Illumina
Hybridization56	RNA96_le1	Illumina
Hybridization57	RNA50_le1	Illumina
Hybridization58	RNA64_le1	Illumina
Hybridization59	RNA93_le1	Illumina
Hybridization60	RNA56_le1	Illumina
Hybridization61	RNA88_le1	Illumina
Hybridization62	RNA67_le1	Illumina
Hybridization63	RNA75_le1	Illumina
Hybridization64	RNA52_le1	Illumina
Hybridization65	RNA95_le1	Illumina
Hybridization66	RNA57_le1	Illumina
Hybridization67	RNA61_le1	Illumina
Hybridization68	RNA87_le1	Illumina
Hybridization69	RNA89_le1	Illumina
Hybridization70	RNA77_le1	Illumina
Hybridization71	RNA69_le1	Illumina
Hybridization72	RNA91_le1	Illumina
Hybridization73	RNA62_le1	Illumina
Hybridization74	RNA54_le1	Illumina
Hybridization75	RNA65_le1	Illumina
Hybridization76	RNA83_le1	Illumina
Hybridization77	RNA74_le1	Illumina
Hybridization78	RNA86_le1	Illumina
Hybridization79	RNA78_le1	Illumina
Hybridization80	RNA70_le1	Illumina
Hybridization81	RNA66_le1	Illumina
Hybridization82	RNA51_le1	Illumina
Hybridization83	RNA63_le1	Illumina
Hybridization84	RNA68_le1	Illumina
Hybridization85	RNA76_le1	Illumina
Hybridization86	RNA80_le1	Illumina
Hybridization87	RNA59_le1	Illumina
Hybridization88	RNA72_le1	Illumina
Hybridization89	RNA84_le1	Illumina
Hybridization90	RNA55_le1	Illumina
Hybridization91	RNA49_le1	Illumina
Hybridization92	RNA71_le1	Illumina
Hybridization93	RNA92_le1	Illumina
Hybridization94	RNA85_le1	Illumina
Hybridization95	RNA94_le1	Illumina

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