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Modified acid guanidinium thiocyanate-phenol-chloroform RNA extraction method which greatly reduces DNA contamination.
| Content Provider | Semantic Scholar |
|---|---|
| Author | Siebert, Paul D. Chenchik, Alex A. |
| Copyright Year | 1993 |
| Abstract | The most common and consistently successful methods for isolating intact RNA are modifications of the Guanidinium Thiocyanate method of Chirgwin et al. (1). One widely used modification of this method, developed by Chomczynski and Sacchi (2), involves extraction of the guanidinium thiocyanate homogenate with phenol—chloroform at reduced pH. The Acid Guanidinium Thiocyanate Phenol -Chloroform (AGPC) method is very rapid and does not require ultra centrifugation making it ideal for isolating RNA from multiple samples. During our study of gene expression in the mouse macrophage cell line, RAW 264.7, by reverse transcriptase-polymerase chain reaction (RT-PCR) we observed that the AGPC method would often yield considerable amounts of genomic DNA contamination. RT—PCR experiments can often tolerate small amounts of DNA contamination by using PCR primers designed to span one or more introns such that efficient PCR amplification is only achieved from cDNA or so that the PCR products derived from genomic DNA and cDNA can be distinguished on an agarose gel. However this is not possible for genes that lack introns or |
| File Format | PDF HTM / HTML |
| DOI | 10.1093/nar/21.8.2019 |
| PubMed reference number | 7684133 |
| Journal | Medline |
| Volume Number | 21 |
| Issue Number | 8 |
| Alternate Webpage(s) | https://oup.silverchair-cdn.com/oup/backfile/Content_public/Journal/nar/21/8/10.1093/nar/21.8.2019/2/21-8-2019.pdf?Expires=1492657318&Key-Pair-Id=APKAIUCZBIA4LVPAVW3Q&Signature=CMd5FYcq0ColhCDXfLryMrPGZr~1y4Krx569hqDTkRoHilJOojycbFm1Umld8D-fwDAOw53TeC90qdYhTEkhRzIEgkAb-1iFiQ0u6ZLJ-684bVS7ezqRFFc7J99YfcXhpoSKvrUMbIuPTXNcUcJSSJ9-sdJYPpOt1FyMBqdGy1d4W1dzzUmvP3UegcfzWTf6T9ErSiEnr2y3YNMX4~FcTx5Jovl2h97Db8x-t5CcMwVmgDK99SvHJ6yQ1U-8-b3Vwp35Wd~9-2ZpS32Aagi6~XM10Qmso5QGlV914eI99rmtXky2fOmLYjxx5y3cIykLllSQak~HR2cRTPAuDVd9QA__ |
| Alternate Webpage(s) | https://doi.org/10.1093/nar%2F21.8.2019 |
| Journal | Nucleic acids research |
| Language | English |
| Access Restriction | Open |
| Content Type | Text |
| Resource Type | Article |
