کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
6490819 | 43387 | 2015 | 19 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Research of methods to detect genomic mutations induced by CRISPR/Cas systems
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کلمات کلیدی
موضوعات مرتبط
مهندسی و علوم پایه
مهندسی شیمی
بیو مهندسی (مهندسی زیستی)
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
The indel-forming non-homologous end joining (NHEJ) pathway repairs double strand breaks in mammalian genomes, resulting in mutation formation following genome editing. Common techniques employed to identify these mutations include the amplified fragment length polymorphism (AFLP) and SURVEYOR assays, which are time consuming, laborious, and only offer a low level of sensitivity. An alternative to these approaches, which is examined in this study, is based on the quantitative PCR high-resolution melting (qPCR-HRM) curve analysis technique and offers simple implementation, is capable of handling large sample sizes, takes no more than 90Â min, and produces sensitive results. Using the newly discovered RNA-guided CRISPR/Cas systems, the IL2RG and EMX1 genes were edited in the human 293T cell line in order to compare the mutation detection accuracies of the aforementioned methods. Genomic mutations were simulated by mixing mutated DNA fragments with normal fragments along a concentration gradient. The results of this comparative study showed that the HRM approach was both reproducible and accurate.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Journal of Biotechnology - Volume 214, 20 November 2015, Pages 128-132
Journal: Journal of Biotechnology - Volume 214, 20 November 2015, Pages 128-132
نویسندگان
Kun Wang, Dong Yi Mei, Qian Nan Liu, Xiao Huan Qiao, Wei Min Ruan, Tian Huang, Geng Sheng Cao,