NCBI Magic-BLAST RNA-seq mapping tool

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First you need to create a BLAST database for your genome or transcriptome. For your reference sequences in a FASTA file, use this command line:

makeblastdb -in <reference.fa> -dbtype nucl -parse_seqids -out <database_name> -title "Database title"

The -parse_seqids option is required to keep the original sequence identifiers. Otherwise makeblastdb will generate its own identifiers, -title is optional.

For more information on makeblastdb see NCBI BLAST+ Command Line User Manual.

Magic-BLAST will work with a genome in a FASTA file, but will be very slow for anything larger than a bacterial genome (about 5 million bases), so we do not recommend it.

Example

To create a BLAST database from the reference file my_reference.fa

$ cat my_reference.fa
>sequence_1 Homo sapiens hemoglobin subunit alpha 2 (HBA2), mRNA
CATAAACCCTGGCGCGCTCGCGGGCCGGCACTCTTCTGGTCCCCACAGACTCAGAGAGAACCCACCATGG
TGCTGTCTCCTGCCGACAAGACCAACGTCAAGGCCGCCTGGGGTAAGGTCGGCGCGCACGCTGGCGAGTA
TGGTGCGGAGGCCCTGGAGAGGATGTTCCTGTCCTTCCCCACCACCAAGACCTACTTCCCGCACTTCGAC
CTGAGCCACGGCTCTGCCCAGGTTAAGGGCCACGGCAAGAAGGTGGCCGACGCGCTGACCAACGCCGTGG
CGCACGTGGACGACATGCCCAACGCGCTGTCCGCCCTGAGCGACCTGCACGCGCACAAGCTTCGGGTGGA
CCCGGTCAACTTCAAGCTCCTAAGCCACTGCCTGCTGGTGACCCTGGCCGCCCACCTCCCCGCCGAGTTC
ACCCCTGCGGTGCACGCCTCCCTGGACAAGTTCCTGGCTTCTGTGAGCACCGTGCTGACCTCCAAATACC
GTTAAGCTGGAGCCTCGGTAGCCGTTCCTCCTGCCCGCTGGGCCTCCCAACGGGCCCTCCTCCCCTCCTT
GCACCGGCCCTTCCTGGTCTTTGAATAAAGTCTGAGTGGGCAGCAAAAAAAAAAAAAAAAAA
>sequence_2 Homo sapiens alpha one globin (HBA1) mRNA, complete cds
CAGACTCAGAGAGAACCCACCATGGTGCTGTCTCCTGCCGACAAGACCAACGTCAAGGCCGCCTGGGGTA
AGGTCGGCGCGCACGCTGGCGAGTATGGTGCGGAGGCCCTGGAGAGGATGTTCCTGTCCTTCCCCACCAC
CAAGACCTACTTCCCGCACTTCGACCTGAGCCACGGCTCTGCCCAGGTTAAGGGCCACGGCAAGAAGGTG
GCCGACGCGCTGACCAACGCCGTGGCGCACGTGGACGACATGCCCAACGCGCTGTCCGCCCTGAGCGACC
TGCACGCGCACAAGCTTCGGGTGGACCCGGTCAACTTCAAGCTCCTAAGCCACTGCCTGCTGGTGACCCT
GGCCGCCCACCTCCCCGCCGAGTTCACCCCTGCGGTGCACGCCTCCCTGGACAAGTTCCTGGCTTCTGTG
AGCACCGTGCTGACCTCCAAATACCGTTAAGCTGGAGCCTCGGTGGCCATGCTTCTTGCCCCTTGGGC

use this command line

makeblastdb -in my_reference.fa -out my_reference -parse_seqids -dbtype nucl

Note that the word following ‘>’ is a sequence identifier that will be used in Magic-BLAST reports. The identifier should be unique.

Download a genome

There are several ways to download whole genomes, transcriptomes, or selected sequences from NCBI.

NCBI Datasets

You can search and download genome and transcript sequences from NCBI Datasets Genome page. Search for an organism, select an assembly, and you will see download options.

For example, you can download the GRCh38 assembly of the human genome, from https://www.ncbi.nlm.nih.gov/datasets/genome/GCF_000001405.40/.

NCBI EDirect tools

To download human chromosome 1 using NCBI EDirect tools use:

search -db nucleotide -query NC_000001 | efetch -format fasta >NC_000001.fa