Bioinformatics- how to perform Global Allignment.

There are three stages of alignment:

1) Scoring matrix

2) Trace back

3) Alignment

1) Scoring matrix:

There are some rules to perform this step:

1) Gap must be placed in the first box.

2) Direction is from zero to high diagonally.

3) Score of each box comes from three places:

• Bottom box (+Gap)  
• Beside box (+Gap) 
• Diagonal box (match/mismatch)

4) Value of arrow added from significant box.

2)Trace back:

In this step we perform following things:

1) First of all find highest value and mark it.

2) Trace highest value back toward zero.

3)Allignment

Traceback arrow direction.

 Let us discuss each step one by one.

1) Scoring matrix:

Choose two sequences , like,       

A T C G

     --- T C G

Score them like, Match = +1,  Mismatch = -1,  Gap = -2

Draw coordinates, and put these sequence on both coordinates, but put gap at the beginning of each coordinate.

• Add  “0”  in first box, and started to add value of gap in Column 1 (C-1) and Row 4 (R-4) by the addition of gap value in the incoming value, like add -2 in the box adjacent to 0, then add value of gap in -2, value would be -4, now add gap value in -4, answer would be -6 and so on, fill C-1 and R-4 in same manner.

• 
• We have to put value in remaining 9 boxes, suppose we have to put value in box shown by arrow, in this box, values can come from three different directions. From the box behind it, from the box bottom it and fro the box which is diagonal, as shown by small arrows.
• For box behind it, add gap value in -2, answer would be -4, for box bottom it add gap value in -2 answer would be -4, and for the value from diagonal we have to look match mismatch base pair, as, T and A are mismatch so put value of mismatch in this box, which is -1. Now, in this box there are 3 values -4, -4 and -1. From all these 3 value put that value in this box which is largest one. As -1 is largest, value in this box would be -1.

• From these 3 arrows consider only one arrow from which actual value is com and remove other two arrows. Continue this process in the same way all boxes appeared would be like as shown.

2)Trace back

• For trace back we have to find out the highest score value, which is +1 in this case.
• From this value we trace back to 0
• Note that this +1 has come from 0 that is diagonal to it. Put arrow from +1 to 0
• Now this zero come from -1 that is diagonal to it as shown by arrow , put arrow in back direction (from 0 to -1)
• Value of -1 again come from diagonal whose value is -2, put arrow from -1 to -2. We have a straight line of trace backing.

3)Alignment

• Now note the arrow. If arrows are horizontal or vertical put gap.
• If arrow is diagonal put  sequence characters.
• Note +1 comes from value which is at diagonal. And its horizontal and vertical characters are G and G. so put  G

                                                                                                 G

• For next corresponding box “0” is come from diagonal value. Its sequence character are C and C again a match. So put sequence character.         

                                                 C       G

                                                 C       G

• Same is for other diagonal value as shown by above table. So sequence would be    

                                             T      C      G

                                             T      C      G

• For last value there is corresponding sequence” A” vertically and a “Gap” horizontally. So sequence would be,          

                                                                               A     T      C     G

                                                                               ---   T      C     G

So, whole sequence have been obtained again.