Identifying what the gene sequence encodes: Select blastx (translated nucleotide to protein). The top hit will identify the organism this DNA came from. Paste the fastA file sequence into the Enter Query Sequence box (or upload the file) and hit the BLAST button. Identifying which organism the DNA sample came from: Select Nucleotide BLAST (nucleotide-nucleotide). This can be performed at the website for the National Center for Biotechnology Information (NCBI) 1. Using BLAST to find homologous sequences: Using BLAST to find homologous sequences is one of the most important and frequently used skills in molecular biology. Make a fasta file by making the following format: >fasta file TTTCTCTCTCT etc etc 5. Generate a fasta sequence file by selecting 'Chromatogram Data', selecting all the sequence (command a), copying (command c) and pasting the sequence into TextEdit (command v). In the standard mode, trim off the bad sequence at the start and end of the sequence by selecting the region and clicking delete (bad sequence refers to regions containing Ns and tall and uneven peaks). Open the sequence file (KB824.ab1) in SnapGene Viewer. Once the dmg file is downloaded, click on it to open the install window and drag the SnapGene Viewer icon to the Applications folder. To look at and edit the chromatogram: Download free SnapGene Viewer 4.1.5 from: Click on download button. Analysing Sanger Sequencing Reactions: Sanger sequencing was performed on a region of the recombinant plasmid. What gene does the sequence show the highest homology to? 4.3) What is the E value and does this indicate that this hit is reliable? Explain.īIOL2343 2021 Assignment 1: Recombinant DNA Technology 4. Questions: 4.1) What organism did the DNA sample come from? 4.2) Include your blastx results. A E value of <10 -5 is considered reliable. * For each hit an E value is reported, defined as the expected number of alignments with scores as good or better than that of the hit in the database by chance. SOLVED: BIOL2343 2021 Assignment 1: Recombinant DNA Technology 4.
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