|
| Status |
Public on Dec 27, 2018 |
| Title |
Positive_control_genomic_changes_due_to_re-replication_at_ChrIV 567 kb_1 |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
negative control MC2A strain in G2/M
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
yjl number: YJL8427
gal induction: n/a
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
DNA was purified by organic extraction and ethanol precipitation. This re-replicating genomic DNA was prepared by Method 1 as described in a previous publication from Joachim Li's lab: Finn and Li JJ PloS Genetics 2013.
|
| Label |
Cy3
|
| Label protocol |
amino-allyl-dUTP incorporation with Klenow. Chemical coupling of dye to aa-dUTP labelled DNA.
|
| |
|
| Channel 2 |
| Source name |
Positive control YJL8363_a MC2A ars317-567kb strain in G2/M induced in galactose for 3 h
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
yjl number: YJL8363
galactose induction: re-replication at ChrIV 567 kb
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
DNA was purified by organic extraction and ethanol precipitation. This re-replicating genomic DNA was prepared by Method 1 as described in a previous publication from Joachim Li's lab: Finn and Li JJ PloS Genetics 2013.
|
| Label |
Cy5
|
| Label protocol |
amino-allyl-dUTP incorporation with Klenow. Chemical coupling of dye to aa-dUTP labelled DNA.
|
| |
|
| |
| Hybridization protocol |
90% of the re-replicating DNA preparation and 1 µg of the reference DNA were labeled, respectively, with Cy3 and Cy5 fluorescent dye (Fisher Scientific 45-001-270). The Cy3 and Cy5 labeled DNA were isolated, combined and applied to an in-house printed microarray. Hybridization was performed in 3X SSC, 25mM HEPES pH7.0, 0.25% SDS at 63oC for 18 to 24 hours.
|
| Scan protocol |
Arrays were then scanned using an Axon Scanner 4B. GenePix Pro6.0 and BatchReplicationAnalyzer4.2 were then used to generate copy number information across the genome.
|
| Description |
Supp. Figure 1
|
| Data processing |
Data were filtered in GenePix to flag as bad features that had (1) obvious defects, (2) saturated pixels, (3) regression r^2 values less than 0.5 or (4) fewer than 55% of their pixels with flourescence intensity greater than 2 standard deviations above background. BLAST analysis was used to generate a filter that would flag as bad elements whose PCR product were excessively repetitive. Raw Cy5/Cy3 median of ratios values were normalized such that the average ratio was equal to 1.
|
| |
|
| Submission date |
Dec 26, 2018 |
| Last update date |
Dec 28, 2018 |
| Contact name |
duyen Bui |
| E-mail(s) |
duyenthanh.bui@ucsf.edu
|
| Phone |
6073793347
|
| Organization name |
UCSF
|
| Lab |
Joachim Li
|
| Street address |
600 16th street
|
| City |
San Francisco |
| State/province |
CALIFORNIA |
| ZIP/Postal code |
94107 |
| Country |
USA |
| |
|
| Platform ID |
GPL3412 |
| Series (1) |
| GSE124382 |
DNA re-replication is susceptible to nucleotide level mutagenesis |
|
