کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
8369281 | 1543043 | 2018 | 6 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Effect of endogenous reference genes on digital PCR assessment of genetically engineered canola events
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کلمات کلیدی
NSFHMG-I/YAcyl-ACP thioesteraseddPCRGMOCruciferindPCRCCFDMFDigital PCR - PCR دیجیتالgenetically modified organism - ارگانیزم اصلاح شده ژنتیکیDNA extraction - استخراج DNAFID - درDroplet digital PCR - قطره PCR دیجیتالgenetically engineered - مهندسی ژنتیکpolymerase chain reaction - واکنش زنجیره ای پلیمرازPCR - واکنش زنجیرهٔ پلیمرازHigh-mobility group protein - پروتئین گروه بالا تحرکSingle nucleotide polymorphism - پلیمورفیسم تک نوکلئوتیدیSNP - چندریختی تک-نوکلئوتیدReference genes - ژنهای مرجعCanola - کانولا
موضوعات مرتبط
علوم زیستی و بیوفناوری
علوم کشاورزی و بیولوژیک
علوم کشاورزی و بیولوژیک (عمومی)
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
Droplet digital PCR (ddPCR) has been used for absolute quantification of genetically engineered (GE) events. Absolute quantification of GE events by duplex ddPCR requires the use of appropriate primers and probes for target and reference gene sequences in order to accurately determine the amount of GE materials. Single copy reference genes are generally preferred for absolute quantification of GE events by ddPCR. Study has not been conducted on a comparison of reference genes for absolute quantification of GE canola events by ddPCR. The suitability of four endogenous reference sequences (HMG-I/Y, FatA(A), CruA and Ccf) for absolute quantification of GE canola events by ddPCR was investigated. The effect of DNA extraction methods and DNA quality on the assessment of reference gene copy numbers was also investigated. ddPCR results were affected by the use of single vs. two copy reference genes. The single copy, FatA(A), reference gene was found to be stable and suitable for absolute quantification of GE canola events by ddPCR. For the copy numbers measured, the HMG-I/Y reference gene was less consistent than FatA(A) reference gene. The expected ddPCR values were underestimated when CruA and Ccf (two copy endogenous Cruciferin sequences) were used because of high number of copies. It is important to make an adjustment if two copy reference genes are used for ddPCR in order to obtain accurate results. On the other hand, real-time quantitative PCR results were not affected by the use of single vs. two copy reference genes.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Biomolecular Detection and Quantification - Volume 15, May 2018, Pages 24-29
Journal: Biomolecular Detection and Quantification - Volume 15, May 2018, Pages 24-29
نویسندگان
Tigst Demeke, Monika Eng,