Computational protocol: Fragmented mitochondrial genomes of the rat lice, Polyplax asiatica and Polyplax spinulosa: intra-genus variation in fragmentation pattern and a possible link between the extent of fragmentation and the length of life cycle

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Protocol publication

[…] Illumina sequence-reads obtained from the mt minichromosomes of Po. asiatica and Po. spinulosa were assembled into contigs with Geneious 6.1.6 []; the parameters for assembly were minimum overlap identity 98% and minimum overlap 50 bp. tRNA genes were identified using tRNAscan-SE [] and ARWEN []. Protein-coding genes and rRNA genes were identified with BLAST searches of GenBank [,]. Identical sequences shared between genes were identified with Wordmatch []. Sequence alignments were with Clustal X []. The size and circular organization of each mt minichromosome of Po. asiatica and Po. spinulosa identified by sequence-read assembly were verified by PCR (Figure B, C). A pair of outbound primers (forward and reverse) was designed from the coding region of each minichromosome (see Additional file ). The two primers in each pair were next to each other with a small gap or no gap in between. PCRs with these primers amplify the full or near full length of each minichromosome if it has a circular organization. PCR set-up, cycling conditions, agarose gel electrophoresis and size measurement were the same as described above. The nucleotide sequences of the mt genomes of Po. asiatica and Po. spinulosa have been deposited in GenBank under accession numbers KF647751–KF647772. […]

Pipeline specifications

Software tools Geneious, tRNAscan-SE, ARWEN, EMBOSS, Clustal W
Application Genome annotation
Organisms Rattus norvegicus, Homo sapiens, Sus scrofa, Bandicota indica, Rattus tanezumi, Pediculus humanus