Research Details
| Research Title | Genomic DNA Yield in Cattle Using Different Methods of Isolation |
| Researcher(s) | Don Carlo Batara |
| Research Category | Project |
| Research Status | completed |
| Duration | Aug 01, 2014 to Aug 31, 2015 |
| Commodity | Livestock |
| Research Site(s) | |
| Source of Fund(s) | |
| Brief Description | This study provides a baseline information of the differences in purity, quantity and cost per nanogram (µl) yield of DNA. High quantity, purity and integrity of nucleic acids from mammalian tissues are essential to molecular biology procedures. This study was carried out to compare the quality, quantity and purity of genomic DNA from whole blood extracted using different protocols and to determine the most cost effective and time efficient method of extraction. Blood samples were collected randomly from the cattle population in Banna, Ilocos Norte. Samples were laid out in CRD with sub-sampling with four replicates for whole blood group and three replicates for blood samples stored in FTA cards. Extracted genomic DNA from whole blood samples and FTA cards obtained from cattle were compared in terms of quantity (concentration of DNA extracted) and quality (260/280 ratio) by LSD. Purity of the samples was verified in 1.2% agarose gel. In whole blood, protocol 1 yields the highest DNA concentration however its purity is below the range while Protocol 3 yielded reasonable amount and quality of DNA. On the other hand, genomic DNA extracted from FTA cards, Protocol 1 yielded the highest concentration of quality DNA. This method could be the useful tool which may apply for further molecular analysis studies using long storing time materials. |
| Expected Output | Many genomic DNA extraction methods have been described for prokaryotes and eukaryotes, from cell sample to specific tissues. However, despite the bulk of molecular studies, few methods has been compared and studies to ensure efficiency of protocols. It is oftentimes experienced that at the middle of a research work, an experiment fails due to contamination of inhibitors in the DNA samples and low concentration is also a factor. Thus, the need to identify methods to obtain intact genomic DNA of good quality and quantity. Based on the results, the following conclusions were drawn: For whole blood samples, 1. When considering the quantity in genomic DNA isolation, Protocols 1, 2 and 3 yields within the acceptable range before down streaming in which the first was significantly higher followed by protocols 3 then 2; 2. Protocols 3 and 4 showed considerable purity which both falls within the acceptable range; 3. In terms of both quantity and quality considered in isolation, Protocol 3 is the most suitable method to use; and 4. The cheapest method of genomic DNA isolation is Protocol 1 followed by 3. Then, protocols 2 and 3 follows. For blood samples stored in FTA cards. 1. Protocol 1 yields significantly higher genomic DNA yield as compared to other Protocols; 2. Protocol 1 yields genomic DNA with considerable purity which is higher than the other protocols; and 3. Protocol 1 is the cheapest method with acceptable quantity and quality of genomic DNA yield. |
| Abstract | High quantity, purity and integrity of nucleic acids from blood are essential to molecular researches such as microsatellites, indexing and genotyping among others. This study was carried out to compare the quality, quantity and purity of genomic DNA from whole blood extracted using different protocols and to determine a cost effective and time efficient method of genomic DNA isolation. Extracted genomic DNA from whole blood samples and FTA cards obtained from cattle were compared in terms of quantity (concentration of DNA extracted) and quality (260/280 ratio) by LSD. In whole blood, protocol 1 yields the highest DNA concentration of 157.38 ng/µl and is above the acceptable range (50 ng/µl) however its purity of 1.55 OD 260/280 is below the desired range (1.8-2 OD 260/280) while Protocol 3 yielded reasonable concentration (82.38 ng/µl) and quality (1.82 OD 260/280) of DNA. For genomic DNA extracted from FTA cards, Protocol 1 yielded the highest concentration (9.10 ng/µl) and purity (1.77 OD 260/280) pf DNA. This study provides a standard procedure for DNA isolation considering the concentration and purity of extracts. It also gives a baseline information of the protocol for FTA cards that could be further explored and optimized to isolate DNA for down streaming. |
