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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Dec 22, 2021 |
| Title |
MT1040 |
| Sample type |
SRA |
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| Source name |
tumor-bearing lung
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| Organism |
Mus musculus |
| Characteristics |
genotype: KrasLSL-G12D;Rosa26LSL-tdTomato;H11LSL-Cas9
Sex: Male
lentivirus_dose: 1.00E+05
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| Growth protocol |
Lung tumors were initiated via intratracheal delivery of 60 µL of a Lenti-sgRNA/Cre pool. Lung tumors were developed for 15 weeks.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Lung tissue was extracted temporarily frozen for later processing. Prior to tissue homogenization and proteinase K digestion, each lung sample was spiked with three cell lines bearing known barcodes (5.00E+05 cells of each cell line). Genomic DNA was then isolated by phenol-chloroform extraction followed by ethanol precipitation from the total lung tissue of each mouse.
One-step PCR targeting lentiviral barcodes to append Illumina adapters, unique dual indices for multiplexing, and six to nine random bases to increase sequence complexity.
Custom Amplicon Library (Tuba-seq)
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NextSeq 500 |
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| Description |
Amplicon library derived from lentiviral barcodes integrated within genomic DNA of the lungs
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| Data processing |
Library strategy: Tuba-seq
Each read is expected to contain an 8-nucleotide sgID region followed by a 30-nucleotide barcode (BC) region (GCNNNNNTANNNNNGCNNNNNTANNNNNGC), and each of the 20 Ns represent random nucleotides with roughly equal representation of A, T, G and C.
The sgID region corresponds to the sgRNA encoded within the Lenti-sgRNA/Cre vector that initiated a given tumor. We require a perfect match between the sequence in the forward read and one of the known sgIDs included within the Lenti-sgRNA/Cre pool.
We require the forward and reverse read to agree completely within the 30 nucleotide sequence to be further processed. Any tumors that are within a Hamming distance of two from a larger tumour is regarded as spurious, as they are likely to result from sequencing or PCR errors, and thus are excluded from downstream analyses.
The tumor size (number of neoplastic cells) is calculated by normalizing the number of reads to the three benchmark spike-in cell lines (5.00E+05 cells for each benchmark cell line) added to each sample prior to lung homogenization/digestion and genomic DNA extraction.
Supplementary_files_format_and_content: Summary of all tumors in the study, including the identity and genotype of the originating mouse of origin, the sgID and clonal identifier, read count, and tumor size.
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| Submission date |
Jul 06, 2021 |
| Last update date |
Dec 22, 2021 |
| Contact name |
Christopher Murray |
| Organization name |
Stanford University
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| Department |
Genetics
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| Lab |
Winslow
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| Street address |
1291 Welch Rd. B261, Beckman Center
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| City |
Stanford |
| State/province |
California |
| ZIP/Postal code |
94305 |
| Country |
USA |
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| Platform ID |
GPL19057 |
| Series (2) |
| GSE179497 |
LKB1 drives stasis and C/EBP-mediated reprogramming to an alveolar type II fate in lung cancer [Primary Tuba-seq] |
| GSE179560 |
LKB1 drives stasis and C/EBP-mediated reprogramming to an alveolar type II fate in lung cancer |
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| Relations |
| BioSample |
SAMN20070140 |
| SRA |
SRX11359210 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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