How to assign allele numbers and sequence type after amplifying housekeeping genes by PCR for genotyping of S. pneumoniae strains by MLST using bioedit ?
after having the sequence for each, ......
Take this example:
let's say that you did mlst for your bacteria, on of genes you amplified is named: ace gene ... you sent for sequencing and and you received the raw forward and reverse sequences for this gene. then you have to trim the sequence to obtain the latest edited sequence....thats means you need to align them with reference sequence .....
go to https://pubmlst.org/ ==> chose your microorganism, ===> download allele===>chose your gene====> copy your reference sequence ==> use allele sequence as a leader to align with your forward and reverse sequences using any bioinformatics software (like bioedit). Then you will get your allele sequence. ....... to know what is the code, go to mlst.net===>locus query ==> single locus ==> past your sequence >>>>>submit ....then record the digit.
Do these steps for the rest six genes ..so that you'll have allelic profile of seven digits
for example: (1,4,3,3,1,7,21) ...... in https://pubmlst.org/, go to profile query ==> allelic profile ==> enter your digits ==> submit ==> then you will get your sequence type (ST).
then contact with https://pubmlst.org/ curator to assign the ST.
Note: sometimes the ST is not presented in the mlst database, then it might be novel ST which is common .......
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