Dear samsmuthu, 

This will not be possible with the help of RegularExpression Validation.

You have to use Custom Validator.

Refer code given below.

HTML

<asp:TextBox runat="server" ID="txtASP"></asp:TextBox>
<asp:CustomValidator runat="server" ClientValidationFunction="Validate" ControlToValidate="txtASP" ErrorMessage="Invalid" />
<script type="text/javascript">
    function Validate(sender, args) {
        var key = document.getElementById(sender.controltovalidate).value;
        var spKeys = new Array();
        spKeys.push("#");
        spKeys.push("%");
        spKeys.push("&");
        spKeys.push("{");
        spKeys.push("}");
        spKeys.push("\\");
        spKeys.push("<");
        spKeys.push(">");
        spKeys.push("*");
        spKeys.push("?");
        spKeys.push("/");
        spKeys.push("$");
        spKeys.push("'");
        spKeys.push(":");
        spKeys.push("@");
        spKeys.push("+");
        spKeys.push("`");
        spKeys.push("|");
        spKeys.push("=");
        spKeys.push("\"");
        if (key.indexOf("  ") == -1) {
            for (var i = 0; i < key.length; i++) {
                if (spKeys.indexOf(key.charAt(i)) == -1) {
                    args.IsValid = true;
                } else {
                    args.IsValid = false;
                    return false;
                }
            }
        } else {
            args.IsValid = false;
            return false;
        }
    }
</script>

Screenshot