The utilization of biotinylated RNA baits on captured sequencing of cancer marker genes functional regions
- VernacularTitle:Хорт хавдрыг нөхцөлдүүлэгч генийн мутацийг РНХ-проб ашиглан илрүүлэх дэвшилтэт арга, технологи
- Author:
Byambasuren B
1
;
Dulamsuren O
2
;
Lkhagvadorj G
3
;
Amarsanaa E
3
;
Khurelbaatar S
3
;
Shiirevnyamba A
1
;
Batsaikhan B
1
;
Zanabazar E
4
Author Information
1. Mongolian National University of Medical Science
2. National Pathology Center
3. National Cancer Center
4. Zanaspex ltd
- Publication Type:Journal Article
- Keywords:
NGS;
cancer marker gene;
targeted sequencing;
capturing;
enrichment;
RNA baits
- From:Mongolian Medical Sciences
2021;198(4):3-14
- CountryMongolia
- Language:Mongolian
-
Abstract:
Background:Nucleic acid sequencing is a multi-step process taken place in medical research or diagnostic
laboratories. Since the emerge of second generation sequencing technology generally referred as
next generation sequencing (NGS), the mass parallel reads covering human genome or transcriptome
is achieved by cost cut down over thousand folds. Though the technology made tremendous push
forward to various applications, its data analysis time and effort still takes worrisome time and human
effort, bringing the emerge of next-step demand: targeted mass sequencing of only desired part
from human genome or transcriptome with lower material cost and labor. By targeted sequencing,
both run cost and data analysis process can be further cut down, and the read results are more
reliable on changes such as determining varied number of repeats, heterozygote alleles, deletions,
chromosomal scale abnormality and more.
Objective:In this study, we explored the utilization of biotinylated RNA baits on captured sequencing of cancer
marker genes functional regions.
Method:Targeted NGS was achieved by capturing desired genomic regions using preparatory nucleic acid
probes. RNA bait capturing of desired genomic regions has shown to have high specificity and quality.
The study was carried out with informed consent obtained from patients, with the approval №53 in
2018.03.15 by Medical Ethics committee, Ministry of Health, Mongolia.
Result:By preparing library of biotinylated RNA baits with 75000 unique sequences, we achieved mass
parallel sequencing of human 410 cancer-marker-genes’ exons and UTRs with average read depth
~760, and covered thousands of SNPs on 5 genomic DNA samples. Tissue samples derived from
breast cancer and ovary cancer had SNP and deletion on 7 marker genes (BRCA1, BRCA2, ATM, BRIP1, PTEN, TP53, RAD51C) not registered in database.
Conclusion:Experiments showed RNA baits with up to 117 nucleotide length, produced from ssDNA oligonucleotide
stock, can be utilized to capture desired regions of human genome, and bring the cost of captured
mass sequencing to 1500 USD, with 93.14-93.33% of Q30 read quality.
- Full text:2021-198(4)-3-14.pdf
