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Date: 18-3-2021
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Date: 16-5-2016
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Date: 4-4-2021
1459
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Nested PCR
Experience shows that whenever PCR is performed on a patient sample, the amplification products very often exhibit a smear in the following ethidium bromide-stained agarose gel electrophoresis. This is due to nonspecific priming in the background of high amounts of host cell nucleic acids and the need to perform a large number of thermocycles (typically 40). Although the first PCR amplification resulted in a smear in the agarose gel, the specific nucleic acids sought will usually be highly enriched compared with the starting material. If a small amount of the first PCR is used for a second PCR amplification with a new set of internal primers, then a well-defined PCR fragment typically results if the infectious agent is present in the initial sample. The use of a second round of PCR with a new set of internal primers is called a nested PCR. If only one internal primer is used for the second round of thermocycling, it is called a semi-nested PCR.
Nested and semi-nested PCR therefore increase the sensitivity as much as 1000-fold compared with conventional PCR9 and increase specificity, but the techniques require more work and longer assay times and carry an increased risk of contamination.
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علامات بسيطة في جسدك قد تنذر بمرض "قاتل"
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أول صور ثلاثية الأبعاد للغدة الزعترية البشرية
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مكتبة أمّ البنين النسويّة تصدر العدد 212 من مجلّة رياض الزهراء (عليها السلام)
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