release 2.2.3 2010-09-17 ===============================================

Minor error corrections, minor cleanup.
This is a stable release.

release 2.2.2 beta 2010-03-04 ==========================================

This release adds a performance enhancement for inputs involving
SEQUENCE_PRIMER_PAIR_OK_REGION_LIST.

This release adds two new tags, PRIMER_MIN_LEFT_THREE_PRIME_DISTANCE
and PRIMER_MIN_RIGHT_THREE_PRIME_DISTANCE

release 2.2.1 alpha 2009-10-28 ==========================================

This release adds several performance enhancements.

release 2.2.0 alpha 2009-10-23 ==========================================

This release adds multiple new features to primer3_core and related
C code.

The enhancements fall under several categories:

1. PRIMER/OLIGO SECONDARY STRUCTURE AND PRIMER-DIMER AND TEMPLATE
MISPRIMING PREDICTION

We added capabilities for predicting secondary structure,
primer-dimer, and template mispriming based on a thermodynamic model.

INPUT/OUTPUT TAGS:

Added tags PRIMER_THERMODYNAMIC_ALIGNMENT,
PRIMER_INTERNAL_MAX_{HAIRPIN,SELF_ANY_TH,SELF_END_TH,TEMPLATE_MISHYB_TH},
PRIMER_INTERNAL_WT_{HAIRPIN,SELF_ANY_TH,SELF_END_TH,TEMPLATE_MISHYB_TH},
PRIMER_MAX_{HAIRPIN,SELF_ANY_TH,SELF_END_TH,TEMPLATE_MISPRIMING_TH},
PRIMER_PAIR_MAX_{COMPL_ANY_TH,COMPL_END_TH,HAIRPIN,TEMPLATE_MISPRIMING_TH},
PRIMER_PAIR_WT_{COMPL_ANY_TH,COMPL_END_TH,HAIRPIN,TEMPLATE_MISPRIMING_TH},
PRIMER_WT_{HAIRPIN,SELF_ANY_TH,SELF_END_TH,TEMPLATE_MISPRIMING_TH}

See the documentation for details. The tag
PRIMER_THERMODYNAMICAL_ALIGNMENT=1 directs primer3_core to use the
thermodynamic alignments.

NEW EXECUTABLE: ntthal

ntthal is analogous to ntdpal.  Between two sequences, ntthal finds
alignment/sec structure, that has the highest melting
temperature. Ntthal is based on nearest-neighbor thermodynamical
approach.  For input parameters to ntthal, execute: ./ntthal (without
arguments).

2. NEW COMMAND LINE FLAGS AND ARGUMENTS

primer3_core now has a more conventional command line.  For example,
you can type

primer3_core -output my_output -error my_error my_input

and primer3_core will take its input from my_input and put
its outputs in the files my_output and my_error.  There
is also a command line flag -echo_settings_file to cause
the settings file contents to be echoed to output.

3. NEW INPUT TAGS TO CONTROL PRIMER LOCATION

See the documentation for SEQUENCE_PRIMER_PAIR_OK_REGION_LIST.  Tags
SEQUENCE_OVERLAP_JUNCTION_LIST,
PRIMER_MIN_3_PRIME_OVERLAP_OF_JUNCTION,
PRIMER_MIN_5_PRIME_OVERLAP_OF_JUNCTION have _replaced_
SEQUENCE_PRIMER_OVERLAP_POS and PRIMER_POS_OVERLAP_TO_END_DIST.  See
the documentation.

4. PERFORMANCE IMPROVEMENTS

release 2.0.0 alpha 2008-12-15 ==========================================

 This is major release, involving enhancements to functionality,
  software architecture, and input and output conventions.

 INPUT/OUTPUT CHANGES:

 * A new command line flag, -io_version=3 provides backward compatible
   behavior for boulder IO input, and boulder IO output that is
   backward compatible except for error messages, warnings, and the
   text of PRIMER...EXPLAIN tags.

 * -io_version=3 IS NOT the default -- you must specify it on the
   command line.
  
 For the new default IO, we renamed many Boulder IO INPUT/OUTPUT TAGS
  for clarification and easier integration into scripts
  
 NEW OR IMPROVED FUNCTIONALITY:
   
 Primer3 'settings' files were added.  A settings file can be read in
  at program start up to set global parameters.  Our intent is that
  parameter values suitable for particular primer design objectives
  can be easily stored and re-used.

 We added new "tasks" such as selection of primer lists, sequencing
  primers, or cloning primers, or the checking of primers or primer pairs
  in the absence of a target sequence. See tag PRIMER_TASK.
 
 It is now possible to request only unique primers when multiple
  primer pairs are requested.  (In the past, each _pair_ was unique,
  but the same primer might occur in more than one pair).  The user
  also has some control over what constitutes a 'unique' primer. See
  tag PRIMER_MIN_THREE_PRIME_DISTANCE.

 When an illegal primer pair is specified for checking primer3, the
  program now assess it.  In the past, primer3 assessed individual
  primers if they violated a constraint, but in some cases not pairs
  that violated constraints on pairs (such as product size).

 We added new functionality with tags 
  SEQUENCE_PRIMER_OVERLAP_POS, PRIMER_POS_OVERLAP_TO_END_DIST,
  PRIMER_MAX_END_GC.  Please see the documentation.

 We added tags PRIMER_PAIR_MAX_COMPL_END and
  PRIMER_PAIR_MAX_COMPL_ANY, which previously took the same values as
  PRIMER_SELF_{END,ANY}

 OTHER CHANGES TO BEHAVIOR:

 We corrected a few errors and a mis-feature in the ways in which
  primer3 searches for primer pairs. Consequently, the latest version
  of primer3 will return different sets of primer pairs in a few
  instances.
  
 INTERNAL CHANGES:

 It is now possible to use the main primer3 functionality from a
  single C function. We cleanly separated the functionality of reading
  in Boulder IO records from the primer choosing / designing
  functionality.  We also substantially simplified the organization
  and documentation of the main primer picking code, now in
  libprimer3.[ch].  The file libprimer3.h specifies the external
  interface, although it is not separately documented. It is also
  possible to create a binary library file that provides this
  functionality (libprimer3.a or libprimer3.so in Unix/Linux).  All C
  code is ANSI C.

release 1.1.2 2007-06-10 ================================================

 Updates to dpal.c so that no error forces an exit.
 Adaptation in primer3_main.c and ntdpal_main.c to 
   updated error handling in dpal().
 Arranged for primer3_main.c align() to check for
   and circumvent local alignments to "targets" < nucleotides
   long, and added tests for this condition.
 Comment and code readability updates in dpal.h and dpal.c

release 1.1.1 3/37/2007 ==================================================

 Minor corrections

release 1.1.0 1/17/2007 ==================================================

 Added tags PRIMER_TM_SANTALUCIA, PRIMER_SALT_CORRECTIONS,
   PRIMER_LOWERCASE_MASKING, PRIMER_DIVALENT_CONC,
   and PRIMER_DNTP_CONC.  See the README for
   details of what these tags do.  Briefly, the first two
   tags allow the use of modern melting temperature 
   calculations.  New tests for this functionality
   were added to the tests/ subdirectory

 NOTE: The oligotm executable now takes additional
   arguments.

 Cleaned up Makefile, and created Makefile for the test
   directory with a target to clean up after tests

 Most (but not all) temporary files created by the
   tests now end in .tmp or .tmp2

 Tried to create more informative test for 
   MAX_PRIMER_LENGTH > DPAL_MAX_ALIGN in primer3_main.c

 NOTE: The oligtotm() function is now provided as
   a library, liboligotm.

 NOTE: non-backward compatible change: removed support of
   flag -2x_compat.

release 1.0.1 11/06 ==================================================

 Identical to 1.0b except for release number

release 1.0b 11/06 ==================================================

  Corrected bug involving check for template mispriming
  Minor code cleanup

release 1.0a   9/06 ==================================================

  No new functionality (bug fixes and minor code cleanup only)

  Bug fixes:

    * Corrected calculation of ..._TEMPLATE_MISPRIMING when
      INCLUDED_REGION was set (thanks to Tomoaki Nishiyama, Advanced
      Sciences Research Center, Kanazawa University, Japan).

    * Corrected error that caused crash when
      PRIMER_WT_TEMPLATE_MISPRIMING was set (thanks to Peng Yu,
      Functional Genomics Group II, Chinese National Human Genome
      Center, Beijing).

    * Added olgiotm_main.c (for executable oligotm)

release 1.0   6/04 ==================================================

  Added ability to search source sequence (template) for
  mispriming or mishybing sites.

  Added _experimental_ facility to calculate Tm on long
  oligos using a GC% based formula.

  Miscellaneous fixes and cleanup, including:

    * Avoiding closing NULL file handles when a mispriming or mishyb
      library is missing

    * Correction to long_seq_tm() in oligotm.c plus addition of
      related tests.

    * Gave user the ability to _not_ treat IUB/IUPAC ambiguity
      codes as a consensus while matching against mispriming
      or mishyb libraries.  Inability to turn off this "feature"
      caused severe problems when libraries contained strings
      of N's:  NNNNNNNNN, since if N is treated as a consensus,
      any base matches.

release 0.9  9/98 ==================================================

  Corrected errors in calculation of amplicon Tm.

release 0.8  6/98 ==================================================

  Added flexible objective function.  Various tags governing
  the penalty weights are not fully documented in this
  release.
  
  Added much changed web interface.

  Added ability to select only a single
  left or right primer or hyb probe.

  Made it possible to continue even if 
  PRIMER_{LEFT,RIGHT,INTERNAL_OLIGO}_INPUT is
  not legal.

  Added PRIMER_{LEFT,RIGHT,INTERNAL_OLIGO}_GC_PERCENT
  output tags.

release 0.7  5/98   ==================================================

  Fixed the 'case problem' for
  PRIMER_{LEFT,RIGHT,INTERNAL_OLIGO}_INPUT (and at the
  same time a related bug).

  Added experimental PRIMER_START_CODON_POSITION tag
  and related outputs.  This feature should still be
  considered experimental and the user should scrutinize
  the output.

  Added PRIMER_PRODUCT_{MAX,MIN}_TM and related outputs.

release 0.6   ==================================================

  Bug fixes (most introduced as part of 0.5 enhancements).

release 0.5   ==================================================

  ** Non-Backward compatible change:
  Fixed inconsistency in output format;
  PRIMER_INTERNAL_n_OLIGO_SEQUENCE
  -> PRIMER_INTERNAL_OLIGO_n_SEQUENCE.
  The old format can be preserved by
  compiling with the precompiler
  symbol USE_OLD_FORMAT_MISTAKE defined.

  Added end-anchored local alignment option to dpal.

  Removed some un-needed dpal implementations.

  Added end-anchored local alignments for checking
  mispriming libraries.

  Added mispriming estimates for primer _pairs_
  (see PRIMER_PAIR_MISPRIMING_LIBRARY),
  with related new output tags.  Trivial change
  in maximum allowable library sequence weight.

  Added an experimental primer position component to
  the objective function (PRIMER_{IN,OUT}SIDE_PENALTY),
  with related new output tags.

  Added sequence quality scores as constraints
  in primer picking (PRIMER_MIN_{,END_}_QUALITY,
  PRIMER_QUALITY_RANGE_{MIN,MAX}) with related
  new output tags.

  Added more error reporting to primer3.cgi when primer3
  exits with a non-0 termination status.

  Fixed bug when non-0, non-1 value was supplied
  for pick internal oligos.

  Added PRIMER_{LEFT,RIGHT,INTERNAL_OLIGO}_INPUT
  for checking or for designing around existing primers or
  internal oligos.

  Added error checking for duplicates of certain
  tags, such as SEQUENCE.

  Added PRIMER_MAX_END_STABILITY and related changes
  to output.

release 0.4b   ==================================================

  Corrected bug in parsing of INCLUDED_REGION tag.

release 0.4   ==================================================

  Added optional check of oligos against "mispriming" or
  "mishyb" libraries  with related changes to formated output
  and to the contents of primer list files.

  Modified search algorithm to compute complementarity/
  similarity measures as late as possible.

  Got rid of quotes (") around error and warning strings.

  Fixed numerical overflow on input sequences longer than
  MAX_SHRT.  (Thanks to Pete Young for finding this one.)

release 0.3   ==================================================

  Changes to dpal to make it substantially faster on the maxgap=1
  case for local and end-anchored complementarity measures.

  Removed restriction on maximum sequence length (by removing
  ftar, fexcl, fexcl_int, fnn, fn, fgc arrays, which necessitated
  internal changes to methods for calculating overlap with
  excluded regions and targets and to calculate spanning of
  targets.

  Modified 'primer list' format (clean up and hopefully got rid
  of spurious regression test discrepancy on some systems due to
  differences in rounding).

  Exit codes for some errors changed.

  Text of error messages for PRIMER_PRODUCT_SIZE -> PRIMER_PRODUCT_SIZE_RANGE.

  Some errors that should have been global (fatal) were not.
  These were fixed and error handling rationalized somewhat.
  
  An error in the display of the start position of targets, and
  excluded regions in formated output was corrected.

  An error in which overlapping product size ranges caused
  primer3 to return duplicate primer pairs was corrected.

  Made regression tests more complete.
  
  Made the maximum number of primer pairs to return a 
  parameter (PRIMER_NUM_RETURN).

  Added optional n-based indexing in input and output
  (PRIMER_FIRST_BASE_INDEX=n).  Has not been tested
  for n < 0.

release 0.2b (10/11/96)  ==================================================
  Added PRIMER_WARNING tag.

  Made it legal for excluded regions and targets to lie outside
  of included region.  Documentation fixes (especially to
  PRIMER_DNA_CONC).

release 0.2a ==================================================
  Tweaks to formated output to make it more complete
  (included complementarity measures);  adjusted tests.

release 0.2 ==================================================
  Made 0-length excluded region legal, and adjusted tests.
  Tweaked error reporting of negative-length excluded regions,
  internal oligo excluded regions, and target.


==============================================================
==============================================================
Compared to non-primer3 progenitors:

Compared to primer 0.5
---------------
Completely different input format.  

It has been reported the 0.5 deleted Ns when they occurred in
primers.  

More stringent self-complementarity defaults.

Primer3 selects internal oligos on request (and produces .int
files if requested).

Compared to both primer 0.5 and primer v2
---------------------------
The format of the contents of .for, .rev (and .int) files is
different.

Primer3 returns a user-specifiable number of primer pairs (or
triples) sorted by "goodness".

Primer3 will find a primer pair if any acceptable pair exists.

Optional n-based indexing into source sequence.

Use of sequence quality and 3' stability as constraints in primer
picking.  Optional positional component to objective function.

Compared to primer v2
-------------
Tag name changes.  However, primer3 should understand most or
all Primer v2 input tags, and should produce v2-compatible output
tag names when the -v2_compat command-line switch is used.

The one exception is that the PRIMER_RECOMMEND tag is no longer
produced. Instead primer3 produces the PRIMER_x_EXPLAIN output
tags.  The format of the data in this tags is different from the
data in v2's PRIMER_RECOMMEND output tag.

Numerous fixes.

Uses the PRIMER_SELF_ANY and PRIMER_SELF_END parameters to govern
maximum allowable complementarity between left and right primers,
as well as complementarity between copies of a single oligo or
within a single oligo.  This behaviour is very close to that of
primer 0.5; self complementarity calculations in v2 were
unreliable.

Primer3 produces much more output information, including the TMs
and self complementarity measures of selected primers.
