BLAST

1. a glimpse under the hood BLAST

2. Prospero: Our revels now are ended. These our actors,As I foretold you, were all spirits, andAre melted into air, into thin air:And like the baseless fabric of this vision,The cloud-capp'd tow'rs, the gorgeous palaces,The solemn temples, the great globe itself,Yea, all which it inherit, shall dissolve,And, like this insubstantial pageant faded,Leave not a rack behind. We are such stuffAs dreams are made on; and our little lifeIs rounded with a sleep. find this

3. Prospero: Our revels now are ended. These our actors,As I foretold you, were all spirits, andAre melted into air, into thin air:And like the baseless fabric of this vision,The cloud-capp'd tow'rs, the gorgeous palaces,The solemn temples, the great globe itself,Yea, all which it inherit, shall dissolve,And, like this insubstantial pageant faded,Leave not a rack behind. We are such stuffAs dreams are made on; and our little lifeIs rounded with a sleep. in here

4. Prospero: Our revels now are ended.These our actors,As I foretold you,were all spirits, andAre melted into air, into thin air:And like the baseless fabric of this vision,The cloud-capp'd tow'rs,the gorgeous palaces,The solemn temples, the great globe itself,Yea, all which it inherit, shall dissolve,And, like this insubstantial pageant faded,Leave not a rack behind. We are such stuffAs dreams aremade on; and our little lifeIs rounded with a sleep.

5. cloud-capp'd tow'rs cloud-capped towers cloudcapp'd towers cloud-tipped towels

6. cloud-capped towers cloud-capp'd tow'rs Our revels now are ended. These our actors, As I foretold you, were all spirits, and Are melted into air, into thin air: And like the baseless fabric of this vision, The cloud-capped towers, the gorgeous palaces, The solemn temples, the great globe itself, Yea, all which it inherit, shall dissolve, And, like this insubstantial pageant faded,

7. cloud-capped towers r: And like the baseless fabric of this vision, The cloud-capped towers, the gorgeous palaces, The solemn temples, the great globe itself, Yea, all which it inherit, shall dissolve, And, like this insubstantial pageant faded,

8. vision, The cloud-capped towers, the gorgeous palaces, The r: And like the baseless fabric of this vision, The cloud-capped towers, the gorgeous palaces, The + + + + + +

9. vision, The cloud-capped towers, the gorgeous palaces, The r: And like the baseless fabric of this vision, The cloud-capped towers, the gorgeous palaces, The solemn temples, the great globe itself, Yea, all which it inherit, shall dissolve, And, like this insubstantial pageant faded, + + + + + +

10. The Natural scenario

12. 1 gene in 1 Panda genome

13. ~ 1 kilobase (kb) in ~ 2.30  gigabases (Gb)

14. in

15. Panda genome GenBank

16. Learning goals [BLAST] Describe the principle Describe the steps Discuss the concepts Use BLAST!

17. Εὑρίσκω Heuristic

18. The algorithm in 13 steps

19. Information reduction in uncertainty degree of surprise

20. Prospero: Our revels now are ended.These our actors,As I foretold you,were all spirits, andAre melted into air, into thin air:And like the baseless fabric of this vision,The cloud-capp'd tow'rs,the gorgeous palaces,The solemn temples, the great globe itself,Yea, all which it inherit, shall dissolve,And, like this insubstantial pageant faded,Leave not a rack behind. We are such stuffAs dreams aremade on; and our little lifeIs rounded with a sleep.

22. vifnknvikctgesqtgntgggqagntggdqagstggspqgstgaspqgstgaspqgstgasqpgssepsnpvssghsvstvsvsqtstssekqdtiqvksallkdymglkvtgpcnenfimflvphiyidvdtedtnielrttlkktnnaisfesnsgslekkkyvklpsngttgeqgsstgtvrgdtepisdssssssssssssssssssssssssssseslpangpdsptvkpprnlqnicetgknfklvvyikent low complexity 1.

23. PAQILWQEDARRKGSM PAQ PAQI nt words = 11 characters aa words = 3 characters PAQIL PAQILW PAQILWQ PAQILWQE PAQILWQED… 2. build word list

24. Biology in BLAST 3. & 4.

25. evaluate matching words e.g. score with BLOSUM PAQ PAQI PAQIL PAQILW QVL PAQILWQ QIA PAQILWQE RIL … PAQILWQED… 3. 20 x 20 x 20 8000 possible scores

26. cloud-capp'd tow'rs cloud-capped towers cloudcapp'd towers cloud-tipped towels QIL QVL QIA RIL …

27. Small P G cyclic side-chain A C Aromatic S - OH T Y Amines W M H N F K E Aliphatic R D Q I V L Charged Hydrophobic Hydrophilic

28. BLOSUM62 8000 words  ca 50 QIL 5 + 4 + 4 T = 10 QVL 5 + 3 + 4 QIA 5 + 4 - 1 RIL … 1 + 4 + 4 4. retain high-scoring words

29. What about nucleotide words?

30. Repeat 3. and 4. for each word listed 5. A 50n polypeptide will yield ~12,500 words

31. create search tree QIL Q N QVL QII Q N NIL V I I QV QI NI 6. L I L L QII QIL QVL NIL

32. plant seeds Query high-scoring words in position n n = 3 QIL,QVL … Exact matches 7. Database

33. stop when maximal score drops by a threshold X + + + + + + 8v1 grow HSPs

34. vision, The cloud-capped towers, the gorgeous palaces, The r: And like the baseless fabric of this vision, The cloud-capped towers, the gorgeous palaces, The + + + + + +

35. vision, The cloud-capped towers, the gorgeous palaces, The The image of “cloud-capped towers”, unites ethereality and might - - + - - - - + - - - -

36. use words at T  longer word list X A X X X Query sequence X X X 8v2 Database sequence grow HSPs

37. extend ‘close’ words as one A X X 8v2 A grow HSPs

38. filter HSPs HSP HSP HSP HSP 9. ≥ S ≤ S S cut-off score

39. determine score significance Could 2 random sequences with the lengths of the query and database resp. score achieve the ≥ HSP score? 10.

40. determine score significance DB S Query S*** S* 10. S**** S** S***** World of randomness S******

41. determine score significance 10. K, λ calculated for the scoring matrix m’ effective length of database seq n’ effective length of query seq

42. determine score significance 10.

43. Extend ≥2 HSPs S 65 40 52 45 Poisson 40 < 45 11. Sum of scores (65+40) > (52+45)

44. Perform Smith-Waterman (local) alignments BLAST BLAST2 12. Calculate bit score (S’) and E-value of each SW alignment

45. Deliver results > E < E 13. E user selected cut-off score

46. A practical example BLAST

47. * Mutation

48. * * Primer binding sites PCR Transposon mutagenesis

49. * * PCR Seq 1 Seq 2

50. Self-study BLAST