xenial (1) melting.1.gz

Provided by: melting_4.3.1+dfsg-2_amd64 bug

NAME

       melting - nearest-neighbor computation of nucleic acid hybridation

SYNOPSIS

       melting [options]

DESCRIPTION

       Melting  computes,  for a nucleic acid duplex, the enthalpy and the entropy of the helix-coil transition,
       and then its melting temperature.  Three types of  hybridisation  are  possible:  DNA/DNA,  DNA/RNA,  and
       RNA/RNA.   The  program  uses the method of nearest-neighbors. The set of thermodynamic parameters can be
       easely changed, for instance following an experimental breakthrough. Melting is a free  program  in  both
       sense  of the term. It comes with no cost and it is open-source. In addition it is coded in ISO C and can
       be compiled on any operating system. Some perl scripts are provided to show how melting can be used as  a
       block to construct more ambitious programs.

OPTIONS

       The options are treated sequentially. If there is a conflict between the value of two options, the latter
       normally erases the former.

       -Afile.nn
              Informs the program to use file.nn as an alternative set of  nearest-neighbor  parameters,  rather
              than  the  default  for  the  specified  hybridisation  type. The standard distribution of melting
              provides some files ready-to-use: all97a.nn (Allawi et al 1997), bre86a.nn (Breslauer et al 1986),
              san96a.nn  (SantaLucia  et  al  1996), sug96a.nn (Sugimoto et al 1996) san04a.nn (Santalucia et al
              2004) (DNA/DNA), fre86a.nn (Freier et al  1986),  xia98a.nn  (Xia  et  al  1998),  (RNA/RNA),  and
              sug95a.nn (Sugimoto et al 1995), (DNA/RNA).

              The  program  will look for the file in a directory specified during the installation. However, if
              an environment variable NN_PATH is defined, melting will search in this one first. Be careful, the
              option -A changes the default parameter set defined by the option -H.

       -Ccomplementary_sequence
              Enters the complementary sequence, from 3' to 5'. This option is mandatory if there are mismatches
              between the two strands. If it is not used, the program will compute it as the complement  of  the
              sequence entered with the option -S.

       -Ddnadnade.nn
              Informs  the  program  to use the file dnadnade.nn to compute the contribution of dangling ends to
              the thermodynamic of helix-coil transition. The dangling ends are not taken into  account  by  the
              approximative mode.

       -Ffactor
              This  is the a correction factor used to modulate the effect of the  nucleic acid concentration in
              the computation of the melting temperature. See section ALGORITHM for details.

       -Gx.xxe-xx
              Magnesium  concentration  (No maximum concentration for the moment). The effect
                 of  ions  on  thermodynamic  stability  of nucleic  acid duplexes is complex,
                 and the correcting functions are  at  best rough  approximations.The published
                 Tm  correction formula for divalent Mg2+ ions of  Owczarzy  et al(2008) can
                 take in account the competitive binding of monovalent and divalent ions on DNA.
                 However this formula is only for DNA duplexes.

       -h     Displays a short help and quit with EXIT_SUCCESS.

       -Hhybridisation_type
              Specifies the hybridisation type. This will set the nearest-neighbor set to use if no  alternative
              set  is  provided  by  the  option  -A (remember the options are read sequentially). Moreover this
              parameter determines the equation to use if the sequence length exceeds the limit  of  application
              of  the  nearest-neighbor approach (arbitrarily set up by the author). Possible values are dnadna,
              dnarna and rnadna (synonymous), and rnarna.  For  reasons  of  compatibility  the  values  of  the
              previous  versions  of melting A,B,C,F,R,S,T,U,W are still available although strongly deprecated.
              Use the option -A to require an alternative set of thermodynamic  parameters.  IMPORTANT:  If  the
              duplex is a DNA/RNA heteroduplex, the sequence of the DNA strand has to be entered with the option
              -S.

       -Iinput_file
              Provides the name of an input file containing the parameters of the run. The input has to  contain
              one  parameter  per line, formatted as in the command line. The order is not important, as well as
              blank lines. example:

              ###beginning###
              -Hdnadna
              -Asug96a.nn
              -SAGCTCGACTC
              -CTCGAGGTGAG
              -N0.2
              -P0.0001
              -v
              -Ksan96a

              ###end###

       -ifile.nn
                Informs  the  program to use file.nn as an alternative set  of  inosine pair
                parameters, rather than  the  default  for the specified hybridisation type.
                The standard distribution of melting provides some  files ready-to-use:  san05a.nn
               (Santalucia et al 2005) for deoxyinosine in DNA duplexes, bre07a.nn (Brent M Znosko
                et al 2007)for inosine in RNA duplexes. Note  that  not all the inosine mismatched
                wobble's pairs have been investigated. Therefore it could be impossible to  compute
                the Tm of a duplex with inosine pairs. Moreover, those inosine pairs are not taken
                into account by the  approximative mode.

       -Ksalt_correction
              Permits one to chose another correction for the concentration in sodium. Currently, one can  chose
              between wet91a, san96a, san98a.  See section ALGORITHM.  TP.  BI. "-k" "x.xxe-xx"
                 Potassium  concentration  (No maximum concentration for the moment). The effect of ions
                 on  thermodynamic  stability  of nucleic  acid duplexes is complex, and the correcting
                 functions are  at  best rough  approximations.The published  Tm  correction formula for
                 sodium ions of Owczarzy et al (2008)is therefore also applicable to buffers containing Tris or
                 KCl. Monovalent K+, Na+, Tris+ ions  stabilize  DNA duplexes
                 with similar potency, and their effects on duplex stability are additive. However this formula
                 is only for DNA duplexes.

       -L     Prints the legal information and quit with EXIT_SUCCESS.

       -Mdnadnamm.nn
              Informs  the  program to use the file dnadnamm.nn to compute the contribution of mismatches to the
              thermodynamic of helix-coil transition. Note that not all the mismatched Crick's pairs  have  been
              investigated. Therefore it could be impossible to compute the Tm of a mismatched duplex. Moreover,
              those mismatches are not taken into account by the approximative mode.

       -Nx.xxe-xx
              Sodium concentration (between 0 and 10 M). The effect of ions on thermodynamic
                stability of nucleic acid duplexes is complex, and the correcting functions
                are at best rough approximations. Moreover, they are generally reliable only
                for [Na+] belonging to [0.1,10M]. If there are no other ions in
                solution, we can use only the sodium correction. In the other case, we use the Owczarzy's
                algorithm.

       -Ooutput_file
              The output is directed to this file instead of the standard output. The name of the  file  can  be
              omitted.  An automatic name is then generated, of the form meltingYYYYMMMDD_HHhMMm.out (of course,
              on POSIX compliant systems, you can emulate  this  with  the  redirection  of  stdout  to  a  file
              constructed with the program date).

       -Px.xxe-xx
              Concentration of the nucleic acid strand in excess (between 0 and 0.1 M).

       -p     Return  the  directory  supposed  to  contain  the  sets  of calorimetric parameters and quit with
              EXIT_SUCCESS. If the environment variable NN_PATH is set, it is  returned.  Otherwise,  the  value
              defined by default during the compilation is returned.

       -q     Turn off the interactive correction of wrongly entered parameter. Useful for run through a server,
              or a batch script. Default is OFF (i.e. interactive on). The switch works in both sens.  Therefore
              if -q has been set in an input file, another -q on the command line will switch the quiet mode OFF
              (same thing if two -q are set on the same command line).

       -Ssequence
              Sequence of one strand of the nucleic acid duplex, entered 5' to 3'. IMPORTANT: If it is a DNA/RNA
              heteroduplex,  the  sequence  of  the  DNA  strand  has  to  be entered. Uridine and thymidine are
              considered as identical. The bases can be upper or lowercase.

       -Txxx  Size threshold before approximative computation. The nearest-neighbour approach will be used  only
              if the length of the sequence is inferior to this threshold.

       -tx.xxe-xx
              Tris buffer  concentration  (No maximum concentration for the moment).
                 The effect  of  ions  on  thermodynamic  stability  of nucleic  acid
                 duplexes is complex, and the correcting functions are  at  best
                 rough  approximations.The published  Tm  correction formula for sodium ions of
                 Owczarzy et al(2008)is therefore also applicable to buffers containing Tris or
                 KCl. Monovalent K+, Na+, Tris+ ions  stabilize  DNA duplexes with similar potency, and
                 their effects on duplex stability are additive. However this formula is only for DNA
                 duplexes. Be careful, the Tris+ ion concentration is about half of the total tris buffer
                 concentration.

       -v     Control the verbose mode, issuing a lot more information about the current run (try it once to see
              if you can get something interesting). Default is OFF. The switch works in both sens. Therefore if
              -v  has been set in an input file, another -v on the command line will switch the verbose mode OFF
              (same thing if two -v are set on the same command line).

       -V     Displays the version number and quit with EXIT_SUCCESS.

       -x     Force the program to compute an approximative tm, based on G+C content. This option has to be used
              with  caution.  Note  that  such  a calcul is increasingly incorrect when the length of the duplex
              decreases. Moreover, it does not take into account nucleic acid concentration, which is  a  strong
              mistake.

ALGORITHM

   Thermodynamics of helix-coil transition of nucleic acid
       The  nearest-neighbor  approach  is  based  on the fact that the helix-coil transition works as a zipper.
       After an initial attachment, the hybridisation propagates laterally.  Therefore, the process  depends  on
       the adjacent nucleotides on each strand (the Crick's pairs).  Two duplexes with the same base pairs could
       have different stabilities, and on the contrary, two duplexes with different sequences but identical sets
       of  Crick's  pairs will have the same thermodynamics properties (see Sugimoto et al. 1994).  This program
       first computes the hybridisation enthalpy and entropy from the  elementary  parameters  of  each  Crick's
       pair.

       DeltaH = deltaH(initiation) + SUM(deltaH(Crick's pair))
       DeltaS = deltaS(initiation) + SUM(deltaS(Crick's pair))

       See  Wetmur  J.G.  (1991) and SantaLucia (1998) for deep reviews on the nucleic acid hybridisation and on
       the different set of nearest-neighbor parameters.

   Effect of mismatches and dangling ends
       The mismatching pairs are also taken into account. However the thermodynamic  parameters  are  still  not
       available  for  every  possible  cases  (notably when both positions are mismatched). In such a case, the
       program, unable to compute any relevant result, will quit with a warning.

       The two first and positions cannot be mismatched. in such a case, the result is  unpredictable,  and  all
       cases are possible. for instance (see Allawi and SanLucia 1997), the duplex

       A          T
        GTGAGCTCAT
        TACTCGAGTG
       T          A

       is more stable than

       AGTGAGCTCATT
       TTACTCGAGTGA

       The dangling ends, that is the umatched terminal nucleotides, can be taken into account.

   Example
       DeltaH(
       AGCGATGAA-
       -CGCTGCTTT
       ) = DeltaH(AG/-C)+DeltaH(A-/TT)
       +DeltaH(initG/C)+DeltaH(initA/T)
       +DeltaH(GC/CG)+DeltaH(CG/GC)+2xDeltaH(GA/CT)+DeltaH(AA/TT)
       +Delta(AT/TG mismatch) +DeltaG(TC/GG mismatch)

       (The same computation is performed for DeltaS)

   The melting temperature
       Then the melting temperature is computed by the following formula:

       Tm = DeltaH / (DeltaS + Rx ln ([nucleic acid]/F))
       Tm in K (for [Na+] = 1 M )
            + f([Na+]) - 273.15
       correction  for  the  salt concentration (if there are only sodium cations in the solution)and to get the
       temperature in degree Celsius.  (In fact some corrections are directly included in the DeltaS see that of
       SanLucia 1998)

   Correction for the concentration of nucleic acid
       If   the  concentration  of  the  two  strands  are  similar,  F  is  1  in  case  of  self-complementary
       oligonucleotides, 4 otherwise. If one strand is in excess (for  instance  in  PCR  experiment),  F  is  2
       (Actually  the  formula  would  have  to  use  the  difference  of  concentrations  rather than the total
       concentration, but if the excess is sufficient, the total concentration can be assumed to be identical to
       the concentration of the strand in excess).

       Note  however,  MELTING makes the assumption of no self-assembly, i.e.  the computation does not take any
       entropic term to correct for self-complementarity.

   Correction for the concentration of salt
       If there are only sodium ions in the solution, we can use the following corrections:

       The correction can be chosen between wet91a, presented in Wetmur 1991 i.e.
       16.6 x log([Na+] / (1 + 0.7 x [Na+])) + 3.85

       san96a presented in SantaLucia et al. 1996 i.e.
       12.5 x log[Na+]

       and san98a presented in SantaLucia 1998 i.e.  a correction of the entropic term without  modification  of
       enthalpy
       DeltaS = DeltaS([Na+]=1M) + 0.368 x (N-1) x ln[Na+]

       Where  N  is  the  length  of  the  duplex  (SantaLucia 1998 actually used 'N' the number of non-terminal
       phosphates, that is effectively equal to our N-1). CAUTION, this correction is meant to  correct  entropy
       values expressed in cal.mol-1.K-1!!!

   Correction  for  the  concentration  of ions when other monovalent ions such as Tris+ and K+ or divalent Mg2+
       ions are added
       If there are only Na+ ions, we can use the correction for the concentration of salt(see  above).  In  the
       opposite  case  ,  we  will  use the magnesium and monovalent ions correction from Owczarzy et al (2008).
       (only for DNA duplexes)

       [Mon+] = [Na+] + [K+] + [Tris+]

       Where [Tris+] = [Tris buffer]/2. (in the option  -t,  it  is  the  Tris  buffer  concentration  which  is
       entered).

       If [Mon+] = 0, the divalent ions are the only ions present
        and the melting temperature is :

       1/Tm(Mg2+)  = 1/Tm(1M Na+) + a - b x ln([Mg2+]) + Fgc x (c + d x ln([Mg2+]) + 1/(2 x (Nbp - 1)) x (- e +f
       x ln([Mg2+]) + g x ln([Mg2+]) x ln([Mg2+]))

       where : a = 3.92/100000.  b = 9.11/1000000.  c = 6.26/100000.  d = 1.42/100000.  e  =  4.82/10000.   f  =
       5.25/10000.  g = 8.31/100000.  Fgc is the fraction of GC base pairs in the sequence and Nbp is the length
       of the sequence (Number of base pairs).

       If [Mon+] > 0, there are several cases because we can have a competitive DNA binding  between  monovalent
       and divalent cations  :

       If  the  ratio  [Mg2+]^(0.5)/[Mon+]  is  inferior to 0.22, monovalent ion influence is dominant, divalent
       cations can be disregarded and the melting temperature is :

       1/Tm(Mg2+) = 1/Tm(1M Na+) + (4.29 x Fgc - 3.95) x 1/100000 x ln([mon+]) + 9.40 x 1/1000000 x ln([Mon+]) x
       ln([Mon+])

       where : Fgc is the fraction of GC base pairs in the sequence.

       If  the  ratio  [Mg2+]^(0.5)/[Mon+]  is  included  in  [0.22,  6[,  we must take in account both Mg2+ and
       monovalent cations concentrations. The melting temperature is :

       1/Tm(Mg2+) = 1/Tm(1M Na+) + a - b x ln([Mg2+]) + Fgc x (c + d x ln([Mg2+]) + 1/(2 x (Nbp - 1)) x (- e + f
       x ln([Mg2+]) + g x ln([Mg2+]) x ln([Mg2+]))

       where : a = 3.92/100000 x (0.843 - 0.352 x [Mon+]0.5 x ln([Mon+])).
               b = 9.11/1000000.       c = 6.26/100000.
               d  =  1.42/100000  x  (1.279  - 4.03/1000 x ln([mon+]) - 8.03/1000 x      ln([mon+] x ln([mon+]).
            e = 4.82/10000.       f = 5.25/10000.       g  =  8.31/100000  x  (0.486  -  0.258  x  ln([mon+])  +
       5.25/1000 x ln([mon+] x ln([mon+] x ln([mon+]).

       Fgc  is  the  fraction  of GC base pairs in the sequence and Nbp is the length of the sequence (Number of
       base pairs).

       Finally, if the ratio  [Mg2+]^(0.5)/[Mon+]  is  superior  to  6,  divalent  ion  influence  is  dominant,
       monovalent cations can be disregarded and the melting temperature is :

       1/Tm(Mg2+) = 1/Tm(1M Na+) + a - b x ln([Mg2+]) + Fgc x (c + d x ln([Mg2+]) + 1/(2 x (Nbp - 1)) x (- e + f
       x ln([Mg2+]) + g x ln([Mg2+]) x ln([Mg2+]))

       where : a = 3.92/100000.  b = 9.11/1000000.  c = 6.26/100000.  d = 1.42/100000.  e  =  4.82/10000.   f  =
       5.25/10000.  g = 8.31/100000.

       Fgc  is  the  fraction  of GC base pairs in the sequence and Nbp is the length of the sequence (Number of
       base pairs).

   Long sequences
       It  is  important  to  realise  that  the  nearest-neighbor  approach  has  been  established  on   small
       oligonucleotides.  Therefore the use of melting in the non-approximative mode is really accurate only for
       relatively short sequences (Although if the sequences are two short, let's say < 6 bp, the  influence  of
       extremities  becomes  too  important  and  the  reliability  decreases  a  lot).  For  long  sequences an
       approximative mode has been designed. This mode is launched if the sequence length  is  higher  than  the
       value given by the option -T (the default threshold is 60 bp).

       The melting temperature is computed by the following formulas:

       DNA/DNA:
       Tm = 81.5+16.6*log10([Na+]/(1+0.7[Na+]))+0.41%GC-500/size

       DNA/RNA:
       Tm = 67+16.6*log10([Na+]/(1.0+0.7[Na+]))+0.8%GC-500/size

       RNA/RNA:
       Tm = 78+16.6*log10([Na+]/(1.0+0.7[Na+]))+0.7%GC-500/size

       This mode is nevertheless strongly disencouraged.

   Miscellaneous comments
       Melting is currently accurate only when the hybridisation is performed at pH 71.

       The  computation  is valid only for the hybridisations performed in aqueous medium.  Therefore the use of
       denaturing agents such as formamide completely invalidates the results.

REFERENCES

       Allawi H.T.,  SantaLucia  J.  (1997).   Thermodynamics  and  NMR  of  internal  G.T  mismatches  in  DNA.
       Biochemistry 36: 10581-10594

       Allawi  H.T.,  SantaLucia  J.  (1998).   Nearest  Neighbor  thermodynamics  parameters  for  internal G.A
       mismatches in DNA.  Biochemistry 37: 2170-2179

       Allawi H.T., SantaLucia J. (1998).  Thermodynamics of internal C.T mismatches in DNA.  Nucleic Acids  Res
       26: 2694-2701.

       Allawi  H.T.,  SantaLucia  J. (1998).  Nearest Neighbor thermodynamics of internal A.C mismatches in DNA:
       sequence dependence and pH effects.  Biochemistry 37: 9435-9444.

       Bommarito S., Peyret N., SantaLucia J. (2000).  Thermodynamic parameters for DNA sequences with  dangling
       ends.  Nucleic Acids Res 28: 1929-1934

       Breslauer  K.J., Frank R., Bl�ker H., Marky L.A. (1986).  Predicting DNA duplex stability from the base
       sequence.  Proc Natl Acad Sci USA 83: 3746-3750

       Freier S.M., Kierzek R., Jaeger J.A., Sugimoto N.,  Caruthers  M.H.,  Neilson  T.,  Turner  D.H.  (1986).
       Improved free-energy parameters for predictions of RNA duplex stability.  Biochemistry 83:9373-9377

       Owczarzy  R., Moreira B.G., You Y., Behlke M.B., Walder J.A.  (2008) Predicting stability of DNA duplexes
       in solutions containing Magnesium and Monovalent Cations. Biochemistry 47: 5336-5353.

       Peyret N., Seneviratne P.A., Allawi H.T., SantaLucia J. (1999).  Nearest Neighbor thermodynamics and  NMR
       of  DNA  sequences  with  internal  A.A,  C.C,  G.G  and  T.T  mismatches.   dependence  and  pH effects.
       Biochemistry 38: 3468-3477

       SantaLucia J. Jr, Allawi  H.T.,  Seneviratne  P.A.  (1996).   Improved  nearest-neighbor  parameters  for
       predicting DNA duplex stability.  Biochemistry 35: 3555-3562

       Sugimoto  N.,  Katoh M., Nakano S., Ohmichi T., Sasaki M. (1994).  RNA/DNA hybrid duplexes with identical
       nearest-neighbor base-pairs hve identical stability.  FEBS Letters 354: 74-78

       Sugimoto N., Nakano S., Katoh M., Matsumura A., Nakamuta H., Ohmichi T., Yoneyama M., Sasaki  M.  (1995).
       Thermodynamic parameters to predict stability of RNA/DNA hybrid duplexes.  Biochemistry 34: 11211-11216

       Sugimoto  N.,  Nakano  S.,  Yoneyama  M.,  Honda  K. (1996).  Improved thermodynamic parameters and helix
       initiation factor to predict stability of DNA duplexes.  Nuc Acids Res 24: 4501-4505

       Watkins N.E., Santalucia J. Jr. (2005). Nearest-neighbor t- hermodynamics of deoxyinosine  pairs  in  DNA
       duplexes. Nucleic Acids Research 33: 6258-6267

       Wright D.J., Rice J.L., Yanker D.M., Znosko B.M. (2007).  Nearest neighbor parameters for inosine-uridine
       pairs in RNA duplexes. Biochemistry 46: 4625-4634

       Xia T., SantaLucia J., Burkard M.E., Kierzek R., Schroeder S.J., Jiao X., Cox  C.,  Turner  D.H.  (1998).
       Thermodynamics  parameters  for  an  expanded  nearest-neighbor  model for formation of RNA duplexes with
       Watson-Crick base pairs.  Biochemistry 37: 14719-14735

       For review see:

       SantaLucia J. (1998) A unified view  of  polymer,  dumbbell,  and  oligonucleotide  DNA  nearest-neighbor
       thermodynamics.  Proc Natl Acad Sci USA 95: 1460-1465

       SantaLucia   J.,  Hicks  Donald  (2004)  The Thermodynamics of DNA structural motifs. Annu. Rev. Biophys.
       Struct. 33: 415 -440

       Wetmur J.G. (1991) DNA probes: applications of the principles of nucleic acid  hybridization.   Crit  Rev
       Biochem Mol Biol 26: 227-259

FILES

       *.nn   Files  containing  the  nearest-neighbor  parameters, enthalpy and entropy, for each Crick's pair.
              They have to be placed  in  a  directory  defined  during  the  compilation  or  targeted  by  the
              environment variable NN_PATH.

       tkmelting.pl
              A  Graphical  User  Interface written in Perl/Tk is available for those who prefer the 'button and
              menu' approach.

       *.pl   Scripts are available to use MELTING iteratively. For instance, the script multi.pl permits one to
              predict  the  Tm  of  several  duplexes  in  one  shot.  The script profil.pl allow an interactive
              computation along a sequence, by sliding a window of specified width.

SEE ALSO

       New versions and related material can  be  found  at  http://www.ebi.ac.uk/compneur/melting/  and  at  at
       https://sourceforge.net/projects/melting/

KNOWN BUGS

       The infiles have to be ended by a blank line because otherwise the last line is not decoded.

       If  an  infile  is called, containing the address of another input file, it does not care of this latter.
       If it is its own address, the program quit (is it a bug or a feature?).

       In interactive mode, a sequence can be entered on several lines with a backslash

       AGCGACGAGCTAGCCTA\
       AGGACCTATACGAC

       If by mistake it is entered as

       AGCGACGAGCTAGCCTA\AGGACCTATACGAC

       The backslash will be considered as an illegal character. Here again, I do not think it is actually a bug
       (even  if  it  is  unlikely, there is a small probability that the backslash could actually be a mistyped
       base).

       Melting is copyright (C) 1997, 2013 by Nicolas Le Novère and Marine Dumousseau

       This program is free software; you can redistribute it and/or modify  it  under  the  terms  of  the  GNU
       General  Public License as published by the Free Software Foundation; either version 2 of the License, or
       (at your option) any later version.

       This program is distributed in the hope that it will be useful, but WITHOUT ANY  WARRANTY;  without  even
       the  implied warranty of MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE.  See the GNU General Public
       License for more details.

       You should have received a copy of the GNU General Public License along with this program; if not,  write
       to the Free Software Foundation, Inc., 59 Temple Place, Suite 330, Boston, MA  02111-1307 USA

ACKNOWLEDGEMENTS

       Nicolas  Joly is an efficient and kind debugger and advisor.  Catherine Letondal wrote the HTML interface
       to melting. Thanks to Nirav Merchant, Taejoon Kwon, Leo Schalkwyk, Mauro Petrillo, Andrew Thompson,  Wong
       Chee  Hong,  Ivano  Zara  for their bug fixes and comments.  Thanks to Richard Owczarzy for his magnesium
       correction. Thanks to Charles Plessy for  the  graphical  interface  files.   Markus  Piotrowski  updated
       TkMELTING  to  cover  version  4.3.  Finally thanks to the usenet helpers, particularly Olivier Dehon and
       Nicolas Chuche.

AUTHORS

       Nicolas Le Novère Babraham Institute, Babraham  Research  Campus  Babraham  CB22  3AT  Cambridge  United-
       Kingdom.  n.lenovere@gmail.com

       Marine  Dumousseau  EMBL-EBI,  Wellcome-Trust  Genome  Campus  Hinxton CB10 1SD Cambridge United-Kingdom.
       marine@ebi.ac.uk

HISTORY

       See the file ChangeLog for the changes of the versions 4 and more recent.