I condensed your script down to the offending part. It still needs some work, but the readline warning disappeared.

#!/usr/bin/perl

use strict;
no strict 'refs';
use warnings;

my $in16S  = '/root/Desktop/pro.faa';

open I16S, '<', $in16S;

my %r_genus16S;
my ($genus16s) = <I16S>;
while ( defined( ${'genus16S'} = readline(*I16S) ) ) {
    if ( $_ =~ />(\w+)\s/ ) {
        ${'genus16S'} = $1;
    }
    if ( exists $${'genus16S'}{ ${'genus16S'} } ) {
        print { 'I16S'; } $_;
        $${'genus16S'}{ ${'genus16S'} } = 'p';
        next;
    }
}

my $no16S = 0;
foreach $_ ( keys %${ 'r_genus16S'; } ) {
    if ( $${'genus16S'}{$_} eq 'n' ) {
        ++$no16S;
        print "$_\n";
        next;
    }
}
close I16S;

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In reply to Re: readline on closed filehandle error by Khen1950fx
in thread readline on closed filehandle error by Oapples

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