I would get the following output from using NCBIStandalone.blastall ?? Code Sniped: BLASTBIN = "/usr/local/bin/blast" BLASTDB = "/usr/local/db/genome/chr19/hs_chr19.fa" blastOutput, errorInfo = NCBIStandalone.blastall(BLASTBIN + "/blastall", 'blastn', BLASTDB, "temp/%s.fasta"% fasta.Accession) Result: <Bio.File.UndoHandle instance at 81121f8> gary