Extracção de DNA

Extracção, Quantificação e Análise de Integridade de DNA
FCT /UNL, DCV, TLBII

2013-2014

Extracção, Quantificação e Análise de Integridade de DNA
FCT /UNL, DCV, TLBII
Catarina Borges, Inês Barroso, Sara Garcia

Abstract
The objective of the following report was the extraction, quantification and analysis of the plasmid DNA in order to know about it’s purity. The extraction was achieved by the alkaline lysis method, the analysis was made by agarose gel 1.2 % electrophoresis and the quantification was made by UV spectrophotometry
(233, 260 and 280 nm). The ratio shown by the spectrophotometry data suggests that the DNA sample was contaminated and even unmanageable due to the fact that the ratio (A260/A280) is less than 1.5.
However our sample was not contaminated with RNA (we used RNase for the purification). The purity results from the UV spectrophotometry showed that A260/A280 was 1.463 and dsDNA concentrations of
4,85 μg/mL.
Despite the affort, the extraction was not efficient because there were some errors during que DNA isolation.
The sample that remained in the well can be genomic DNA. Nevertheless we can not make sure of that using the methods in this process. It remains uncertain.
Results from electrophoresis showed that this process didn’t occur in the way it should, it wasn’t effective enough, tahre was contamination with RNA in some samples. Nevertheless we were able to visualize the three types of DNA ( relaxed, linear and super-coiled)

Sumário
Com o objectivo de extrair, quantificar e analisar a integridade e grau de pureza de amostras de DNA plasmídico utilizou-se o método da lise alcalina, para a extracção, e a análise foi feita por corrida de electroforese em gel de agarose 1,2%, complementada por quantificação por espectofotometria em UV a
230, 260 e 280 nm. Os dados da espectrofotometria acerca desta razão obtidos pelo nosso grupo sugerem que a amostra de DNA se encontra contaminada, podendo mesmo dizer que a