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This article mainly explains "how to use fastANI". Friends who are interested might as well take a look. The method introduced in this paper is simple, fast and practical. Now let the editor take you to learn how to use fastANI.

In comparative genome analysis, we often need to analyze the evolutionary relationships between different genomes, for example, we can use marker proteins to construct phylogenetic trees. In order to make a quantitative comparison, we can also calculate the similarity or evolutionary distance between different genomes for species classification, phylogenetic comparison and so on. Average nucleotide similarity (Average Nucleotide Identity,ANI) is an index to compare the genetic relationship between two genomes at nucleotide level. ANI is defined as the average base similarity between two homologous segments of the microbial genome, which is characterized by a high degree of differentiation among related species.

FastANI is a fast tool for calculating genome-wide ANI, which supports pairwise comparisons among one-to-one, one-to-many and many-to-many genomes. He divides the query sequence into short sequence fragments and uses the MinHash-based sequence mapping engine Mashmap to calculate homologous mappings and estimate consistency. Because it uses a non-comparison method, the calculation speed is greatly improved, but the accuracy is not much different from that of the blast-based method.

In a recent study by Nature communications, the author used fastANI to analyze 90, 000 genomes and found that there is an obvious ANI dividing line between intraspecific and interspecific in most pedigrees. The genomic ANI of the same species is less than 95%, and the genomic ANI of different species is more than 95%. Therefore, 95% ANI is often used as the standard of species division and species clustering.

FastANI can be used by downloading the package from GitHub and decompressing it as follows:

FastANI-Q genome1.fa-r genome2.fa-o output.txtfastANI-Q genome1.fa-- rl genome_list.txt-o output.txt-r,-- ref: reference genomic nucleotide sequence, you can try fasta/fastq and its gzip compressed file-- rl,-- refList: a file containing a list of reference genomes, thus allowing multiple reference genomes-Q,-- query: query genomic nucleotide sequences You can try fasta/fastq and its gzip compressed file-- ql,-- queryList: contains files that query the list of genomes, thus allowing multiple query genomes-k,-- kmer: the kmer size of the alignment cannot be greater than 16. The default is 16murt,-- threads: the number of cores used to run the program. Default is 1--fragLen: fragment length, default is 3000--minFrag: the shortest matching fragment. Default is 50--visualize: output alignment image, only applicable to one-to-one alignment, default off-matrix: output ANI value as the lower triangular matrix, suitable for many-to-many alignment, default off-o,-- output: output file name because most genes in the bacterial genome are about 1000bp, so usually set the fragment length to 1000, for viruses and other small genomes, you can set a smaller fragment length. The one-to-one analysis of the two genomes is as follows: fastANI-Q 951_armatimo.fasta-r 391_armatimo.fasta-o output1.txt-- fragLen 1000

The results are as follows:

Its ANI is 74.7. 2570 is all the sequence segments of the reference genome, and 981 is the homologous fragment on the alignment in the query genome. the ANI value of too few fragments is meaningless and can be removed.

Multiple genomes are compared with each other as follows:

FastANI-ql Armatimonadetes.txt-rl Armatimonadetes.txt-o output2.txt-fragLen 1000-t 10-matrix

The result of the generated matrix is as follows:

The above matrix can be illustrated in R, as follows:

At this point, I believe you have a deeper understanding of "how to use fastANI". You might as well do it in practice. Here is the website, more related content can enter the relevant channels to inquire, follow us, continue to learn!

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