|
| Status |
Public on Jun 03, 2019 |
| Title |
aCGH_HIVneg_DLBCL_pt30 |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
DLBCL
|
| Organism |
Homo sapiens |
| Characteristics |
subject id: pt30
tissue preparation: FFPE
specimen site: Soft tissue chest wall
hiv status: HIV negative
Sex: Female
age: 20
|
| Treatment protocol |
One 5mm and two 10mm sections were cut from each FFPE tissue block. H&E staining was performed on the 5mm section. H&E stained sections were reviewed by a hematopathologist to confirm diagnosis and assess tumor content (tumors with <70% were macrodissected)
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Qiagen Allprep DNA/RNA FFPE kit (cat#80204) was used to extract total mRNA and DNA from FFPE sections as per manufacturer guidelines found here. No more than two 10mm sections were applied to a single spin column.
|
| Label |
Cy-3
|
| Label protocol |
Extracted DNA was treated with DNAse 1 prior to Klenow-based labeling. High molecular weight templates were digested for 30 minutes while the smaller fragmented FFPE-derived DNA samples were digested for only 1 minute. In each case 1 ul of 10x DNase 1 reaction buffer and 2 μl of DNase 1 dilution buffer were added to 7 μl of DNA sample and incubated at room temperature then transferred to 70°C for 30 minutes to deactivate DNase 1. Sample and reference templates were then labeled with Cy-3 dUTP and Cy-5 dUTP respectively using a BioPrime labeling kit (Invitrogen, Carlsbad, CA) according to our published protocols [PMID 26317899]. All labeling reactions were assessed using a Nanodrop assay (Nanodrop, Wilmington, DE) prior to mixing and hybridization to CGH arrays (Agilent Technologies, Santa Clara, CA).
|
| |
|
| Channel 2 |
| Source name |
Reference pooled 46XX
|
| Organism |
Homo sapiens |
| Characteristics |
sample type: Normal female genome
|
| Treatment protocol |
One 5mm and two 10mm sections were cut from each FFPE tissue block. H&E staining was performed on the 5mm section. H&E stained sections were reviewed by a hematopathologist to confirm diagnosis and assess tumor content (tumors with <70% were macrodissected)
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Qiagen Allprep DNA/RNA FFPE kit (cat#80204) was used to extract total mRNA and DNA from FFPE sections as per manufacturer guidelines found here. No more than two 10mm sections were applied to a single spin column.
|
| Label |
Cy-5
|
| Label protocol |
Extracted DNA was treated with DNAse 1 prior to Klenow-based labeling. High molecular weight templates were digested for 30 minutes while the smaller fragmented FFPE-derived DNA samples were digested for only 1 minute. In each case 1 ul of 10x DNase 1 reaction buffer and 2 μl of DNase 1 dilution buffer were added to 7 μl of DNA sample and incubated at room temperature then transferred to 70°C for 30 minutes to deactivate DNase 1. Sample and reference templates were then labeled with Cy-3 dUTP and Cy-5 dUTP respectively using a BioPrime labeling kit (Invitrogen, Carlsbad, CA) according to our published protocols [PMID 26317899]. All labeling reactions were assessed using a Nanodrop assay (Nanodrop, Wilmington, DE) prior to mixing and hybridization to CGH arrays (Agilent Technologies, Santa Clara, CA).
|
| |
|
| |
| Hybridization protocol |
Labeled DNA was hybridized without ozone scavenger and chips washed according to manufacture's protocol for 40 hours in a rotating 65°C oven.
|
| Scan protocol |
All microarray slides were scanned using an Agilent 2565C DNA scanner and the images were analyzed with Agilent Feature Extraction version 11.0 using default settings.
|
| Data processing |
Data was extracted from the TIFF files using Agilent FE 10. The aCGH data was assessed with a series of QC metrics then analyzed using an aberration detection algorithm (ADM2) [PMID16597236]. The latter identifies all aberrant intervals in a given sample with consistently high or low log ratios based on the statistical score derived from the average normalized log ratios of all probes in the genomic interval multiplied by the square root of the number of these probes. This score represents the deviation of the average of the normalized log ratios from its expected value of zero and is proportional to the height h (absolute average log ratio) of the genomic interval, and to the square root of the number of probes in the interval.
|
| |
|
| Submission date |
Sep 05, 2018 |
| Last update date |
Jun 03, 2019 |
| Contact name |
Alanna Maguire |
| E-mail(s) |
maguire.alanna@mayo.edu
|
| Organization name |
Mayo Clinic
|
| Department |
Research
|
| Lab |
CRB 1-250A
|
| Street address |
13400 East Shea Blvd
|
| City |
Scottsdale |
| State/province |
Arizona |
| ZIP/Postal code |
85259 |
| Country |
USA |
| |
|
| Platform ID |
GPL19387 |
| Series (2) |
| GSE119536 |
Enhanced DNA repair and genomic stability identify a novel HIV related Diffuse Large B-cell Lymphoma subtype [aCGH] |
| GSE119537 |
Enhanced DNA repair and genomic stability identify a novel HIV related Diffuse Large B-cell Lymphoma subtype |
|
