BTW, in case my last reply wasn't clear, here's what I was talking about:

#!/usr/bin/perl

use strict;
use warnings;

$/ = undef;     # slurp-mode for input, just in case
while ( <> ) {  # reads stdin or all file names in ARGV
    s/\s+//g;   # remove whitespace
    my $content = $_;  # keep a working copy
    tr/ACGTacgt//d;    # remove all acgt

    if ( length() ) { # anything left?
        print "$ARGV bad content: $_\n";
        do_something_with_bad_data( $ARGV, $content );
    } else {
        print "$ARGV all clean!\n";
        do_something_with_good_data( $ARGV, $content );
    }
}

sub do_something_with_bad_data
{
    my ( $filename, $data ) = @_;
# . . . fix it?  report it to someone?
}

sub do_something_with_good_data
{
    my ( $filename, $data ) = @_;
# . . . whatever you want to do
}
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As a side note, there are some commonly agreed best practices in the Perl community. Among them:

• use the use warnings; pragma rather than the -w flag
• Use lexical filehandles rather than bareword filehandles
• Use the three-argument syntax for the open function

#!/usr/bin/perl

use strict;
use warnings;

my $infile = shift;

open my $INPUT_FILE, "<", $infile" or die "can't open $infile: $!";

local $/; # the whole file will be slurped, even if it has several lin
+es
my $dna = <$INPUT_FILE>;         
if ( $dna  =~ m/[^actg\s]/i ) {
    print "File contains something besides actg sequence.\n";
} else {
    print "good!\n";
}            
close $INPUT_FILE;
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if ( <INPUT_FILE> =~ m/[^actgACTG\s]/ ) {
    ...

Are there any potential problems with this code?

The character class  \s includes  ' ' (space, 0x20) and IIRC  \t \n \r \f other whitespace characters. Your test allows the string read from the file to have any number of any combination of these characters. Please see perlrecharclass.

I must say that I don't understand your desparate, last-ditch efforts to avoid the use of chomp, for it seems very likely that the line you're reading from your file is newline-terminated (whatever a newline happens to be in your OS). Here's how I might handle the file-read-and-validate portion of your program (untested):

use warnings;
use strict;

die "no filename given" unless @ARGV;
my $filename = $ARGV[0];

open my $fh_input, '<', $filename or die "opening '$filename': $!";
my @lines = <$fh_input>;
die "no lines read from '$filename': $!" unless @lines;
close $fh_input or die "closing '$filename': $!";

chomp @lines;
die "more than one line in '$filename'" unless @lines == 1;
my $line = $lines[0];
die "'$filename' contains something other than ACTG sequence"
    if $line =~ m{ [^actgACTG] }xms;

my $result = do_something_with($line);
print "result is: 'result'";

exit;

sub do_something_with { ... }
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