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Mutation detection of TFAM gene by T7EI cleavage assay and sequence analysis.

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posted on 07.03.2019, 18:48 by Vanessa Cristina de Oliveira, Gabriel Sassarão Alves Moreira, Fabiana Fernandes Bressan, Clésio Gomes Mariano Junior, Kelly Cristine Santos Roballo, Marine Charpentier, Jean-Paul Concordet, Flávio Vieira Meirelles, Carlos Eduardo Ambrósio

In the T7EI assay note mutations in numbers 1 to 3. It’s possible to see 2 bands in the center of the gel and the numbers 4, 5, and 6 are controls of clones 1–3. In the PCR sequencing analysis note A, B and C (nucleotides 163–165) showing the PAM region of the sequence. The cleavage site is 3bp prior to PAM (160–162). In A, note the insertion of two A residues at the site that should be GC. At nucleotide 155, there also occurred an insertion of G, normally an A. In B, note the insertion of an A substitution for G. In C the insertion of AT nucleotides involving positions 156 and 157. This site is normally GC. Also insertion of a T at the A site of nucleotide 151, and insertion of C at the G site at nucleotide 144.

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