A Field Guide. GenBank Records. A Typical GenBank Record GenBank Record: Feature Table. The Flatfile Format. part 2. Header. Feature Table.

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A Field Guide part 2 GenBank Records The Flatfile Format Header Feature Table Sequence A Typical GenBank Record GenBank Record: Feature Table LOCUS NM_019570

1 A Field Guide part 2 GenBank Records The Flatfile Format Header Feature Table Sequence A Typical GenBank Record GenBank Record: Feature Table LOCUS NM_ bp mrna linear ROD 28-OCT-2004 DEFINITION Mus musculus REV1-like(S. cerevisiae)(rev1l),mrna ACCESSION NM_ VERSION NM_ GI: KEYWORDS. 1

2 GenBank Record: Feature Table, con t. GenBank Record: sequence Indexing for Nucleotide UID Global Entrez Search: HFE 56!""#"#$%&& ' ()* +,- ' ##./ %"""4.""" ) $""01"$1#& $"".1"#4$"".1"/ ) 2

3 Entrez Nucleotide: HFE 137 records Smarter Query hfe[title] AND human[orgn] 42 records hfe[title] Not HFE Curated HFE splice variants (11 total) hfe[title] AND human[orgn] (con t) Primary data Preview/Index Gateway to Advanced Searches 3

4 Preview/Index Preview/Index: Properties, srcdb 2 2 Properties srcdb Preview/Index: Properties, srcdb Preview/Index: Properties, srcdb AND srcdb refseq[properties] AND srcdb ddbj/embl/genbank[properties] 4

Properties Search Field Properties Search Field: biomol #1 hfe 137 #2 hfe[title] AND human[orgn] 42 #3 #2 AND srcdb refseq[prop] 11 #4 #2 AND srcdb ddbj/embl/genbank[prop] 31 #5 #4 AND gbdiv

5 Properties Search Field Properties Search Field: biomol #1 hfe 137 #2 hfe[title] AND human[orgn] 42 #3 #2 AND srcdb refseq[prop] 11 #4 #2 AND srcdb ddbj/embl/genbank[prop] 31 #5 #4 AND gbdiv pri[prop] 29 #4 #4 AND gbdiv est[prop] #1 hfe 116 #2 hfe[title] AND human[orgn] 42 #3 #2 AND biomol mrna[prop] 29 #4 #2 AND biomol genomic[prop] 13 9 :, 2 :, 2 More Queries 2;) 6 < +3+)8 3 4 srcdb refseq reviewed[prop] AND transcript[title] AND variant[title] More Queries 2;) 6 < +3+)8 3 4 srcdb refseq reviewed[prop] AND transcript[title] AND variant[title] 6 <9 ), 4 topoisomerase[gene name] AND archaea[organism] 9 $= 2[chromosome],: human[organism] AND gene omim [filter] 2 4 integral to plasma membrane [gene ontology] AND cancer[dis] 5

Other Entrez Databases >94 3 +, rat[organism] NOT 0[mrna

2[Chromosome]) AND human[orgn] 9 9 ' ' 5 9 + >884 9 ) #$

8 4 )2 2$? bacteria[organism] AND kinase AND 000.00:002.

6 Other Entrez Databases >94 3 +, rat[organism] NOT 0[mrna count] 874 $ microsat[snp Class] AND 1[Map Weight] AND 2[Chromosome]) AND human[orgn] 9 9 ' ' >884 9 ) #$ Genethon[Map Name] AND human[organism] AND 12[chromosome] 8 4 )2 2$? bacteria[organism] AND kinase AND :002.00[resolution] 6 <9 Gen Biol: Gen Resources Genomic Biology 6

7 Map Viewer Genome Annotation Updates Gen Biol: Gen Resources Genome Projects: microb Genome Projects: microb #%6 ",2B $ 7 ;## 7

8 Gen Biol: Gen Resources Gen Biol: Gen Resources Gen Biol: Gen Resources ' 5 6 <9 8

mouse and paralogs A mouse B chick B frog B Homologene Cluster MLH1 Cluster

9 Homologene No longer orthologs UniGene based orthologs paralogs Protein similarities first Guided by taxonomic tree frog Includes A chick orthologs A mouse and paralogs A mouse B chick B frog B Homologene Cluster MLH1 Cluster A-chain gene B-chain gene gene duplication early globin gene Rice Homolog ' 5 6 <9 9

10 List View Mouse MapViewer: Mouse ADAR MapViewer: Mouse ADAR, 28 Hits 10

11 Mouse MapViewer: Gene Filter MV Hs ADAR exon 3 UTR Maps & Options MapViewer ;;8 ;;,2, 2 '68!A A A A '!:, 9 9 A6+ 7 +) ;;A A ;; ;;9 ;; 9 C!96 ;;+5;; C1 +A;+5 C;D,A 5 + = SNP +)8+, 9 11

12 Maps & Options 8 +,:,+ 5,:,+,:, ' 5 6 <9 12

13 13

Basic Local Alignment Search Tool Outline

important example oligo search BLAST Web

14 Basic Local Alignment Search Tool Outline Web BLAST pre-computed results how BLAST works words; scoring matrices; statistics specialized BLAST algorithms what s new, or important example oligo search BLAST Web Searches, 2006 BLAST Web Searches, 2005 $"""""1 $"""""1 14

15 4 +8 'E, ' >9 5!" E 15

16 Best hits # 3D structures CDD-Search $ %%&' $( &)' $)( ( &' * 8# 8$ 8# 8$ +( 92 E,)- #- 2)F G) $- 82) %- 6 2 B > )(587* B 9( 2* B 9( 2; * 16

17 -.(()/0 112 ATGCTGCTAGTCGATGACGTAGCTA ATGCTGCTAGT TGCTGCTAGTC GCTGCTAGTCG... - AIEKCYTGCTLAQEADDTA AIE IEK LEK, IDK, IQK, IER, IDR, etc EKC () KCY CYT (%()% ()/%*%0 A 4 R -1 5 N D C Q E G H I L K M F P S T W Y V X A R N D C Q E G H I L K M F P S T W Y V X 2!#3.45" 78978:!;11" 78;1< 78; ,=7* Nucleotide: one exact match ATGCTGCTAGTCGATGACGTAGCTA TAGTCGATGA Protein: two matches within 40 residues PHAIEKCYTGCTLAQEADDTA IDK EAD 17

18 !*" Query: IETVYAAYLPKNTHPFLYLSLEISPQNVDVNVHPTKHEVHFLHEESILEV YLS 15 HFL 18 YLT 12 HFV 15 YVS 12 HFS 14 2 YIT 10 HWL 13 etc NFL 13 2 DFL 12 HWV 10 (;)*## etc Query 1 IETVYAAYLPKNTHPFLYLSLEISPQNVDVNVHPTKHEVHFLHEESI 47 +E YA YL K F+ YLSL +SP+ +DVNVHP+K V HFL+++ I Sbjct 287 LEETYAKYLHKGASYFVYLSLNMSPEQLDVNVHPSKRIVHFLYDQEI 333 : ;)) )) 5 B Query 1 IETVYAAYLPKNTHPFLYLSLEISPQNVDVNVHPTKHEVHFLHEESI 47 +E YA YL K F+ YLSL +SP+ +DVNVHP+K V HFL+++ I Sbjct 287 LEETYAKYLHKGASYFVYLSLNMSPEQLDVNVHPSKRIVHFLYDQEI Query 1 IETVYAAYLPKNTHPFLYLSLEISPQNVDVNVHPTKHEVHFLHEESI-LEV 50 +E YA YL K F+YLSL +SP+ +DVNVHP+K VHFL+++ I + + Sbjct 287 LEETYAKYLHKGASYFVYLSLNMSPEQLDVNVHPSKRIVHFLYDQEIATSI ; (587* ;HH HH ))3) (2* 2%"2$"2 2" #0 2 * A G C T A G C T ; #;;% CAGGTAGCAAGCTTGCATGTCA raw score = 19-(6+7)* = 6* CACGTAGCAAGCTTG-GTGTCA I I B90 90B6$ 18

19 A 4 R -1 5 N >! "#!" D C Q E G H I L K M F P S T W Y V X A R N D C Q E G H I L K M F P S T W Y V X 2!#3.45" 2 Position Independent Matrices PAM Matrices (Percent Accepted Mutation)?(@ $ $.1 $.5AB) BLOSUM Matrices (BLOck SUbstitution Matrices) $?(@ ( $ 7*() $ #3.452 * Position-Specific Score Matrix Position-Specific Score Matrix :,;# # 0 J K K A R N D C Q E G H I L K M F P S T W Y V 435 K E S N K P A M A H R D I K S K N I M V K N D L

20 7*C Local Alignment Statistics )2)2N 8 6L ;λ8 6$ ;8M L) λ ) λ8m 2 (λ8; L*1$ 2 6 ( * More info: The Statistics of Sequence Similarity Scores 7 *!" 000 ) ; 7* An alignment BLAST cannot make: 1 GAATATATGAAGACCAAGATTGCAGTCCTGCTGGCCTGAACCACGCTATTCTTGCTGTTG 1 GAGTGTACGATGAGCCCGAGTGTAGCAGTGAAGATCTGGACCACGGTGTACTCGTTGTCG GTTACGGAACCGAGAATGGTAAAGACTACTGGATCATTAAGAACTCCTGGGGAGCCAGTT GCTATGGTGTTAAGGGTGGGAAGAAGTACTGGCTCGTCAAGAACAGCTGGGCTGAATCCT GGGGTGAACAAGGTTATTTCAGGCTTGCTCGTGGTAAAAAC GGGGAGACCAAGGCTACATCCTTATGTCCCGTGACAACAAC 9 *D(0 20

. Score Score 84 = 290 290 bits bits (741), (741), Expect Expect O'E,8H = 7e-77 7e-77 Identities Identities = 147/331 147/331 (44%), (44%), Positives Positives = 206/331 206/331 (61%), (61%), Gaps

21 . Score Score 84 = bits bits (741), (741), Expect Expect O'E,8H = 7e-77 7e-77 Identities Identities = 147/ /331 (44%), (44%), Positives Positives = 206/ /331 (61%), (61%), Gaps Gaps = 8/331 8/331 (2%) (2%) Frame Frame = 'E,8$8(2*4 # Megablast Discontiguous Megablast PSI-BLAST PHI-BLAST.(9 E +?.( $? $ + $ $ F(@ Uses discontiguous word matches Better for cross-species comparisons WORD SIZE default minimum blastn 11 7 megablast

?- W = 11, t = 16, coding: 1101101101101101 W = 11, t = 16, non-coding: 1110010110110111 W = 12, t = 16, coding: 1111101101101101 W = 12, t = 16, non-coding: 1110110110110111 W = 11, t = 18, coding:

22 ?- W = 11, t = 16, coding: W = 11, t = 16, non-coding: W = 12, t = 16, coding: W = 12, t = 16, non-coding: W = 11, t = 18, coding: W = 11, t = 18, non-coding: W = 12, t = 18, coding: W = 12, t = 18, non-coding: W = 11, t = 21, coding: W = 11, t = 21, non-coding: W = 12, t = 21, coding: W = 12, t = 21, non-coding: W <OQ t +) 4 ' PE ) 3 -') $""$O#/(%*4KK";0?! 2".?-# 22

23 .().()?0.(7*000 7*9. Query: NM_ Drosophila melanogaster CG18582-PA (mbt) mrna, (3244 bp) /note= mushroom bodies tiny; synonyms: Pak2, STE20, dpak2 Database: nr (nt), Mammalia[orgn] MegaBLAST = poor hits Discontiguous megablast = numerous hits... 23

24 7*9? *9<64. 2!"#$ % &'( ")*+,-. /-' ( 01 24

F; F%E 78;'E,84A 'E,84A )# R$ New sp O52692 MTS1_STRCS Modification methylase ScaI (N-4 cytosin... 108 2e-23 + New sp P29538 MTH1_HAEPA Modification methylase HpaI (Adenine-spe.

8 3e-18 + New sp Q45971 MTC1_CAUCR Modification methylase CcrMI (Adenine-sp... 83.5 5e-16 + New sp O30569 MTS1_RHIME Modification methylase SmeIP (Adenine-sp... 81.

7 3e-14 + Gene New sp Q04845 MTC1_CITFR Modification methylase CfrBI (N-4 cytosi... 67.3 4e-11 + New sp P30774 MTX1_XANCC Modification methylase XcyI (N-4 cytosin... 65.

9 4e-10 New sp P09358 MTD22_STRPN Modification methylase DpnIIB (Adenine-... 61.6 2e-09 New sp P23941 MTB1_BACAM Modification methylase BamHI (N-4 cytosi... 60.

.. 54.6 2e-07 + New sp P43871 MTH3_HAEIN Modification methylase HindIII (Adenine-... 53.9 4e-07 +)2 New sp P18051 MTB2_BACAM Modification methylase BamHII (N-4 cytos... 53.1 6e-07 + New sp O52513 MTS1_STRFI Modification methylase SfiI (N-4 cytosin.

25 F; F%E 78;'E,84A 'E,84A )# R$ New sp O52692 MTS1_STRCS Modification methylase ScaI (N-4 cytosin e-23 + New sp P29538 MTH1_HAEPA Modification methylase HpaI (Adenine-spe e-22 New sp P23192 MTM2_MORBO Modification methylase MboII (Adenine-sp e-20 + New sp P28638 YHDJ_ECOLI Hypothetical adenine-specific methylase yhd e-18 + New sp Q45971 MTC1_CAUCR Modification methylase CcrMI (Adenine-sp e-16 + New sp O30569 MTS1_RHIME Modification methylase SmeIP (Adenine-sp e-15 + New sp P20590 MTH1_HAEIN Modification methylase HinfI (Adenine-sp e-15 + New sp Q2YMK2 MTB1_BRUA2 Modification methylase BabI (Adenine-spe e-14 + Gene New sp Q04845 MTC1_CITFR Modification methylase CfrBI (N-4 cytosi e-11 + New sp P30774 MTX1_XANCC Modification methylase XcyI (N-4 cytosin e-10 + New sp P14243 MTC9_CITFR Modification methylase Cfr9I (N-4 cytosi e-10 + New sp P50178 MTL22_LACLC Modification methylase LlaDCHIB (Adenin e-10 New sp P09358 MTD22_STRPN Modification methylase DpnIIB (Adenine e-09 New sp P23941 MTB1_BACAM Modification methylase BamHI (N-4 cytosi e-09 New sp Q58893 MTM5_METJA Modification methylase MjaV (N-4 cytosin e-08 + New sp P14230 MTSM_SERMA Modification methylase SmaI (N-4 cytosin e-08 New sp P34721 MT1B_MORBO Modification methylase MboIB (Adenine-sp e-07 + New sp P43871 MTH3_HAEIN Modification methylase HindIII (Adenine e-07 +)2 New sp P18051 MTB2_BACAM Modification methylase BamHII (N-4 cytos e-07 + New sp O52513 MTS1_STRFI Modification methylase SfiI (N-4 cytosin e-06 New sp Q58392 MTM1_METJA Modification methylase MjaI (N-4 cytosin e-06 + New sp Q9S4X2 YUBD_ECOLI Putative methylase yubd e-06 New sp O68556 MTB1_BACSU Modification methylase BglI (N-4 cytosin e-05 + New sp O59647 MTMW_METWO Modification methylase MwoI (N-4 cytosin e-04 + New sp P14827 MTEC_ENTCL Modification methylase EcaI (Adenine-spe e-04 New sp P71366 T3MH_HAEIN Putative type III restriction-modificati New sp P40814 T3MO_SALTY Type III restriction-modification system New sp P08763 T3MO_BPP1 Type III restriction-modification system New sp Q9LAI2 MTB1_BACSQ )2 Modification methylase BslI (N-4 2 cytosin )2 2 2 sp P25076 CY11_SOLTU Cytochrome c1 heme protein, mitochondria sp Q83DD0 SYV_COXBU Valyl-tRNA synthetase (Valine--tRNA ligase)

-2-5 -4-1 -1-2 -5-3 -2 267 D -1-1 -1 5-1 0 2-1 0-4 -2 3 1-5 -2 0-1 -6-5 -4 268 L -3 4-1 -1-2 -1-1 -5 1 1 0 2-2 -3-4 0 2-2 -1 1 269 V -1-5 -6-6 -1-5 -5-5 -6 3 1-5 1-1 -5-4 -3-5 -1 6 270 V -1-5 -6-6

-5 5-2 -5-4 -5-1 5-6 -4-3 -3 3-5 0-5 -4-5 9-4 -5-2 -5-3 -3-5 -1-3 -1 0 2-5 -5-2 C 0-3 -3-3 9 275 G -2-5 -3-4 -5-4 -4 7-4 -6-6 -3-5 -5-4 -2-3 -4-5 -5 Q -1 1 0 0-3 5 E -1 0 0 2-4 2 5 G 0-2 0-1 -3-2 -2

26 G. #3.45 Last position-specific scoring matrix computed, weighted observed perc A R N D C Q E G H I L K M F P S T W Y V 265 P D D L V V D A 4 R I N D F 274 G C G Q E G H I L K M F P S T W Y V X G274, A R N D C Q E G H I L K M F P S T W Y V X G275 BLOSUM62 5( Basic Local Alignment Search Tool 23 &4?( S ( * S B 9':3 B A B!H!+)8 S S )! B 9883 B!H!+! S )! B A!!9! S B 68:3 S B 593 S 2 B 88:3 S 2) B 7,:3 S B ) 3 S 2 B ) S 3! B 3 26

27 Protein BLAST Databases New Nucleotide Databases Protein nr traditional GenBank records refseq = NP_, XP_ swissprot pdb pat env_nr nr = nr )F #9=F 8 8 ) 27

28 New Output View New Output View 7 ( +'6 ( ' (8 (8 Sorting Results Sorting Hits: by Score 7( +(( (9 * ((&9 28

Sorting Hits: by Query Start # ) = 6 6 6 <T = < 2 2 ) ) 3 E = (G9 * 4 2 2 = =,3 B#"""" ;3$""" ;2$""" ;#"""" #"""";3$""" 3 Example: Mapping Oligos Onto a Genome Map Oligos

29 Sorting Hits: by Query Start # ) = <T = < 2 2 ) ) 3 E = (G9 * = =,3 B#"""" ;3$""" ;2$""" ;#"""" #"""";3$""" 3 Example: Mapping Oligos Onto a Genome Map Oligos Onto Genome >forward CCATGGCGACCCTGGAAAAGC >reverse CAGCAGCGGCTGTGCCTGCGG U U U >CCATGGCGACCCTGGAAAAGCNNNNNNNNNNCAGCAGCGGCTGTGCCTGCGG forward primer reverse primer -W 7 e

30 Genome BLAST Results Primer Alignments reverse primer forward primer MapViewer MapViewer 30

Sequence View (sv) forward Service Addresses S'E,8 blast-help@ncbi.nlm.

31 Sequence View (sv) forward Service Addresses S'E,8 S9 5 info@ncbi.nlm.nih.gov SC matten@ncbi.nlm.nih.gov reverse 31

BLAST. Varieties of BLAST

BLAST. Varieties of BLAST BLAST Basic Local Alignment Search Tool (1990) Altschul, Gish, Miller, Myers, & Lipman Uses short-cuts or heuristics to improve search speed Like speed-reading, does not examine every nucleotide of database