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Open AcceResearch articleDevelopment of a physical map of the soybean pathogen Fusarium virguliforme based on synteny with Fusarium graminearum genomic DNAJeffry L Shultz*1, Sikander Ali2, Linda Ballard3 and David A Lightfoot4

Address: 1Currently, USDA-ARS, Crop Genetics and Production Research Unit, PO Box 345, Stoneville, MS, 38776, USA, 2Institute of Industrial Biotechnology, GC University Lahore, Pakistan, 3Mississippi Valley State University, Itta Bena, MS. Currently, USDA-ARS MSA Genomics Laboratory, P.O. Box 345, Stoneville, MS, 38776, USA and 4Dept. of Plant, Soil and Agricultural Systems, Genomics and Biotechnology Core Facility, Center for Excellence in Soybean Research, Southern Illinois University, Carbondale IL, 62901, USA

Email: Jeffry L Shultz* - jlshultz@hotmail.com; Sikander Ali - alisbiotech@yahoo.com; Linda Ballard - max@tecinfo.com; David A Lightfoot - ga4082@siu.edu

* Corresponding author

AbstractBackground: Reference genome sequences within the major taxa can be used to assist thedevelopment of genomic tools for related organisms. A major constraint in the use of thesesequenced and annotated genomes is divergent evolution. Divergence of organisms from acommon ancestor may have occurred millions of years ago, leading to apparently un-related andun-syntenic genomes when sequence alignment is attempted.

Results: A series of programs were written to prepare 36 Mbp of Fusarium graminearum sequencein 19 scaffolds as a reference genome. Exactly 4,152 Bacterial artificial chromosome (BAC) endsequences from 2,178 large-insert Fusarium virguliforme clones were tested against this sequence. Atotal of 94 maps of F. graminearum sequence scaffolds, annotated exonic fragments and associatedF. virguliforme sequences resulted.

Conclusion: Developed here was a technique that allowed the comparison of genomes based onsmall, 15 bp regions of shared identity. The main power of this method lay in its ability to aligndiverged sequences. This work is unique in that discontinuous sequences were used for the analysisand information not readily apparent, such as match direction, are presented. The 94 maps andJAVA programs are freely available on the Web and by request.

BackgroundThe soybean pathogen Fusarium virguliforme [1-3] (exFusarium solani f.sp. glycines) causes soybean sudden deathsyndrome (SDS). SDS has caused significant yield lossesin soybean since it was first noted in 1980 [4]. F. virguli-forme is a member of the Martiella section, a Fusariumplant disease complex that contains many related patho-gens of the worlds major crops [5,6]. Fusarium virguliforme

is a hemibiotrophic plant pathogen with a wide hostrange [1,5]. The perfect stage is unknown and all extantstrains appear to be of a single mating type [7]. Tools foranalysis of this economically important genome havebecome a high priority, with two BAC libraries and a col-lection of BAC-end sequences already developed [8-12].These BAC-end sequences (BES) produce two reads fromeach discrete clone, and present 1,000–1,600 base pairs of

Published: 3 August 2007

BMC Genomics 2007, 8:262 doi:10.1186/1471-2164-8-262

Received: 9 February 2007Accepted: 3 August 2007

This article is available from: http://www.biomedcentral.com/1471-2164/8/262

© 2007 Shultz et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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genomic DNA for analysis. A map of ordered, joinedsequences can result if sufficient BACs are end-sequenced[13].

A reference genome is defined here as a genome to whichphenotypic and/or genotypic traits of other genomes maybe compared. The comparison of genotypic traits betweenboth related and distant organisms relies on the informa-tion available for the reference genome. This informationmay be as broad as marker order and karyotype and asspecific as single nucleotide polymorphisms (SNPs). Thedecision on which reference genome to use when severalare available favors that most closely correlated at bothgeneral and specific layers with the organism under study.

Millions of years of divergent evolution may cause prob-lems when comparing distantly related genomes. Evenclosely related species may share sequence homology,conserved synteny and genome architecture among only asmall set of genes and regions after the effects of inver-sions, replications, translocations, insertions, deletions,duplications and a myriad number of other genetic muta-tions. Evolutionary pressure can also cause varyinggenome size and karyotype divergence.

Two genomes were available for use as reference sequencewithin the Fusarium plant pathogen complex, Giberellazeae (F. graminearum) [14] and Nectria haematococca (F.solani f.sp. pisi) [15]. The cereal crop fungal pathogenFusarium graminearum had a nearly complete genomesequence [14], a physical map [16] had the same chromo-some number and shared many gene expression patternswith the pathogenic F. virguliforme [17] (Yuan and Light-foot, unpublished).

The objective was to cluster and align discontinuous BAC-end sequences from F. virguliforme in silico, using sequenceresources available for F. graminearum. Three steps wouldbe taken. First, a method for grouping thousands of dis-crete DNA fragments from an uncharacterized genomebased on the reference genome was developed. Second,the reference genome was annotated for genic (predictedexon) content. Finally, the random BES matches to the ref-erence genome were presented in a linear map format,based on proximity of sequence identities from both endsof each BAC. After these steps, a prediction of regionsspanned by the BAC clones could be made and displayedin an informative manner.

MethodsSequences used for genome comparisonIn June, 2007, there were 4,152 F. virguliforme BES availa-ble on Genbank [8,9]. The BES encoded 2,976,557 bp ofFusarium virguliforme DNA, at an average of 717 bp perrecord and an overall GC content of 53.9%. The

36,093,143 bp of Fusarium graminearum reference genomeis represented in 511 sequence fragments located within19 large (>20 Kbp) and 24 small (<20 Kbp) scaffolds thatcorrespond to 4 chromosomes [16] and have a GC con-tent of 48.4%.

Sequence processingThe creation of an efficient comparison utility required amulti-program process (Figure 1). First, FASTA formattedF. graminearum sequence was processed to remove linefeeds and headings. Second, the file was scanned for 7 bpsequences that contained all four bases (A, T, G and C).Motifs with all four bases were written to an array com-posed of 1,020 possible combinations of the first 5 basepairs (5 base repeats of A, T, G, or C were excluded fromthe analysis).

The array of sequence motifs and positions was loadedinto a second program, followed by the nucleotide-onlyreference sequence. Fasta formatted BES was input inseries, with each BES screened against the F. graminearumgenomic sequence. In brief, a five base nucleotidesequence from the BES was used to access the array ofsequences, allowing the program to attempt to align onlythose sequences that shared the first five base pairs of each15 bp fragment. In addition, the reverse complement ofeach BES was also tested. Execution times for comparisonof 4,152 BES against 36 Mbp of F. graminearum genomicDNA were less than three hours. Output from the compar-ison program was in the form of the reference genomesequence location, BES ID and matching sequence loca-tion. If the BES match was from the reverse complementof a sequence a "-" notation was made.

The identification of exonic fragments was carried outusing TimeLogic® GeneDetective™ (Active Motif Inc.,Carlsbad, CA) In order to directly relate the exons to theassociated BES fragments and to accommodate theprocessing restrictions of Gene Detective, the identical 36Mbp single sequence used for sequence comparison wasdivided into smaller, 999 bp linear order fragments, thentested these against the non-redundant fungal EST data-base (downloaded from NCBI on Nov 16, 2006; 886,100fungal ESTs). We used only the top two ranked hits foreach sequence and removed overlapping results.

A Windows XP o/s, w/2 GHz processor and 512 MB RAMwas used for analyses (other than Gene Detective) per-formed in this experiment. A map using the 19 largesequence Fusarium graminearum scaffolds as a frameworkwas prepared in MapChart [18], with matching F. virguli-forme clones and exon content of F. graminearum. Verifica-tion of alignments was performed using theBLAST2Sequence utility at NCBI [19].

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Program flow diagram for sequence comparison using 36 Mbp of Fusarium graminearum scaffold sequence as a reference for 4,152 Fusarium virguliforme BAC end sequencesFigure 1Program flow diagram for sequence comparison using 36 Mbp of Fusarium graminearum scaffold sequence as a reference for 4,152 Fusarium virguliforme BAC end sequences. All JAVA code used for this comparison is freely available from the authors.

Remove formatting from F.graminearum scaffold

sequences and write as a single, un-interrupted

sequence

Create position array file, filtering repetitive DNA

Load F. graminearum singleline sequence and array into

memory

Load characters 1-15 AATTGCATTGATCTG

Access F. graminearumsequence array with first 5

lettersAATTG 123 894 1893 4294

Use array file to “skip” along F. graminearum DNA

123 894 1893 4294 ___________________________________________

Reverse string and repeat scan to

identify all plus / minus positives

Ouput matches to separate text file

If end of sequence, read

next record

If BAC-end has more characters, advance string

one base

Read F. virguliforme BAC-end sequence recordAATTGCATTGATCTGAATTAA……….

BMC Genomics 2007, 8:262 http://www.biomedcentral.com/1471-2164/8/262

Results and discussionFusarium graminearum scaffold-based mapTo expand the use of reference genomes, sequence testingtechniques must allow for variations in genetic drift andbe capable of reporting associations with varying degreesof certainty [20-22]. In order to meet both of thesedemands, fifteen base pair “windows” were taken fromBAC-end sequences and checked against the referencegenome. In addition, each window was reversed andchecked against the reference, allowing for change in readdirection of BES or of the reference sequence. A fifteenbase pair fragment represents a probability of randomassociation every 1 Bbp (415). Smaller base pair probeswill result in an exponential increase in reported associa-tions. For example, reducing the base pair window to 10would result in a probability of random association every1 Mbp (410), with loss of accuracy as probability dictatesthe likelihood of a random match in the referencegenome.

As Figure 1 indicates, five bp sequence arrays were used toposition the F. virguliforme BAC-end probe sequenceagainst the F. graminearum scaffold sequence.

These arrays performed two vital tasks. First, theyincreased processing speed by nearly 1000× over non-arrayed comparisons. The second advantage related tohow the array was prepared. Each 5 bp sequence was writ-ten to the array only after the next two bases were added,then this 7 bp fragment was tested for the inclusion of all4 nucleotides (A, T, G and C). This provided an initialscreen of low complexity DNA. It was found that this com-plexity test was too stringent when tested on 5 bp frag-ments, and too lax at 8 bp.

Identifying conserved regions between F. virguliformeBACs and F graminearum DNA led to the detection of atotal of 46,590 15 bp matches. These matches allowed theintegration of F. virguliforme BAC-end sequence into thelinear order scaffold map of F graminearum.

There were two criteria built into this experiment: First, a15 bp match had to occur; Second, another 15 bp matchfrom the other end of the clone was required within 200Kbp on the reference sequence. Clones meeting these twocriteria were separated into three groups, based on theexpected bidirectional nature of BAC end sequence datafrom the same clone and the unidirectional nature of theF. graminearum scaffolds. The first group were those thathad alternating match direction between the two BACends and were contained within the same scaffold subse-quence. The second group was identical to group 1, exceptthat the two ends span at least two scaffold subsequences.The third group were those that had unidirectional

matches between the two BAC ends, regardless of scaffoldsubsequence position.

A total of 2,560 putatively syntenic associations wereidentified, encompassing 247,012,452 bp of F. gramine-arum DNA (6.8× genome coverage). A total of 1,339 F. vir-guliforme clones were identified that exhibited plus andminus strands matching up to 200,000 bp apart. The totalDNA covered by +/- alignments was 127,482,241 bp.These clones were separated into 531 that fit within a sin-gle scaffold subsequence (30,915,996 bp; red clones, Fig-ure 2) and 808 that span more than one subsequence(96,566,245 bp; green clones, Figure 2) In addition, 1,221clones exhibited plus and plus or minus and minusstrands matching up to 200,000 bp apart are indicated onthe map in blue. The total DNA covered by +/+ or -/-matches was 119,530,211 bp.

As an example, the match of clone 2M11 (gi #s 38262078and 38262180) with F. graminearum scaffold 1.52 isshown in the sequence alignment panel of Figure 2. The 5'match was in two fragments, (181/230 bp, 1e-35; 95/119bp, 8e-15) around bp position 3,347,394 (3.347 on themap). The 3' match was reversed, with 554/683 bp and anexpect value of 3e-150 around bp position 3,418,000(3.418 on the map). Of additional interest was the loca-tion of perfectly preserved HindIII restriction sites, 92bases upstream (3,347,302 bp) in the 5' F. graminearumsequence and another at the final position (3,418,308 bp)of the 3' match (Figure 2, sequence alignment panel). Thestrength of the sequence matches identified and the nearlyperfect correlation with HindIII sites used for cloning theBAC library provide strong evidence of the effectiveness ofthis sequence correlation technique.

At least 22,614 unique exons encompassing a total of6,888,863 bp were detected in the F. graminearum scaffoldsequences. The match direction of these sequences is indi-cated with red (forward) or magenta (reverse) text in themap. A total of 94 maps (spanning ~400 kb each) weregenerated by this procedure and are available as a self-extracting file [see Additional file 1], which includes asearchable excel file. Each map is viewable as a completelyscalable enhanced metafile using the Microsoft™ Pictureand Fax Viewer program.

Future workThis report is important because it describes the use of F.graminearum sequence as a reference to develop genomeresources for F. virguliforme. The conserved syntenyinferred will greatly accelerate the genomic analysis of F.virguliforme. The map may also accelerate the discovery ofpathogenic gene clusters [16]. Parallels in gene expressionunderlying the pathogenic mechanisms of both F. virguli-forme and F. graminearum may be identified [17]. Strate-

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Graphical representation of predicted sequence alignments between Fusarium graminearum reference sequence and Fusarium virguliforme BAC-end sequencesFigure 2Graphical representation of predicted sequence alignments between Fusarium graminearum reference sequence and Fusarium virguliforme BAC-end sequences. Inset key panel shows how color of predicted match was determined. A match from both ends of the BAC within 200 kb was required for display. Uni-directional matches are indicated in blue. Reversed order matches are indicated in green if they cross a scaffold marker (indicating a discontinuous sequence) and red if they are entirely within a sin-gle, continuous scaffold. Fusarium graminearum exonic fragments are indicated in red (5'-3') and magenta (3'–5'). Inset sequence alignment panel illustrates the strength of the match between both ends of clone “2M11” and scaffold 1.52 (1e-35 top and 3e-150, bottom) The presence of HindIII restriction fragments was verified in the F. graminearum sequence, as would be predicted by the position of 2M11.

4_g1c10fs.r1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_g1c10fs_5',_mRNA_sequen BI189734_g1c10fs.r1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_g1c10fs_5',_mRNA_sequen 320.3BI189733_g1c10fs.f1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_g1c10fs_3',_mRNA_sequen BU068840_2568_F11_K22ZT5_Mature_peri thecia_Gibberella_zeae_cDNA,_mRNA_sequence 320.5

BU062571_Fgr_1_P12_T7_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 320.8DR670610_EST1060727_FvO_Gibberella_moniliformis_cDNA_clone_FVOBC43,_mRNA_sequence DR673221_EST1063338_FvO_Gibberella_moniliformis_cDNA_clone_FVOBT38,_mRNA_sequence 321.2

697_i3g12fs.r1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_i3g12fs_5',_mRNA_sequen BI190697_i3g12fs.r1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_i3g12fs_5',_mRNA_sequen 321.4DR681576_EST1071693_FvO_Gibberella_moniliformis_cDNA_clone_FVODR36,_mRNA_sequence 321.6

DR681577_EST1071694_FvO_Gibberella_moniliformis_cDNA_clone_FVODR36,_mRNA_sequence EC596533_FU071f01_Asexual_conidial_life-cycle_of_Fusarium_oxysporum_f._sp._cucumerinum_Fusarium_oxysporum_f._sp._cucumerinum_cDNA_clone_FU07DR647177_EST1037294_FvN_Gibberella_moniliformis_cDNA_clone_FVNA837,_mRNA_sequence DR619024_EST1009152_FvH_Gibberella_moniliformis_cDNA_clone_FVHDM49,_mRNA_sequence 321.7

Scaffold1.48 DR647177_EST1037294_FvN_Gibberella_moni liformis_cDNA_clone_FVNA837,_mRNA_sequenceDR638225_EST1028850_FvM_Gibberella_moniliformis_cDNA_clone_FVMB631,_mRNA_sequence DR638225_EST1028850_FvM_Gibberella_moniliformis_cDNA_clone_FVMB631,_mRNA_sequence 321.8

DR647178_EST1037295_FvN_Gibberella_moniliformis_cDNA_clone_FVNA837,_mRNA_sequence BU067870_1631_C01_F01ZT5_Mature_peri thecia_Gibberella_zeae_cDNA,_mRNA_sequenceBU067870_1631_C01_F01ZT5_Mature_perithecia_Gibberella_zeae_cDNA,_mRNA_sequence EC593220_FU013d03_Asexual_conidial_life-cycle_of_Fusarium_oxysporum_f._sp._cucumerinum_Fusarium_oxysporum_f._sp._cucumerinum_cDNA_clone_FU01 321.9

BU060319_Fgr-C_0_P20_T3_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence BU060319_Fgr-C_0_P20_T3_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequenceBU060319_Fgr-C_0_P20_T3_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 322.1

BU060320_Fgr-C_0_P20_T7_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence BU060320_Fgr-C_0_P20_T7_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequenceDT527759_AP1C6_Subtracted_cDNA_library_from_Fusarium_graminearum_infected_wheat_heads_(sFHB)_Gibberella_zeae_cDNA_clone_AP1C6_5',_mRNA_seque 322.2

EB530979_Fg13_03o24_T3_Fg13_AAFC_ECORC_Fusarium_graminearum_mycelium_partial_cDNA_Gibberella_zeae_cDNA_clone_Fg13_03o24,_mRNA_sequence 322.4T3_Fg13_AAFC_ECORC_Fusarium_graminearum_mycelium_partial_cDNA_Gibberella_zeae_cDNA_clone_Fg13_03o24,_mRNA_sequence EB530671_Fg13_02k18_T3_Fg13_AAFC_ECORC_Fusarium_graminearum_mycelium_partial_cDNA_Gibberella_zeae_cDNA_clone_Fg13_02k18,_mRNA_sequence

DR628130_EST1018258_FvI_Gibberella_moniliformis_cDNA_clone_FVICO81,_mRNA_sequence DR617918_EST1008046_FvH_Gibberella_moniliformis_cDNA_clone_FVHCY46,_mRNA_sequence 322.5

DR618802_EST1008930_FvH_Gibberella_moniliformis_cDNA_clone_FVHDJ77,_mRNA_sequence BE605250_e4g01fs.f1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_e4g01fs_3',_mRNA_sequen 322.6DR668941_EST1059058_FvO_Gibberella_moniliformis_cDNA_clone_FVOB127,_mRNA_sequence DR668942_EST1059059_FvO_Gibberella_moniliformis_cDNA_clone_FVOB127,_mRNA_sequence 322.7

0_c4c07fs.f1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_l ibrary_Fusarium_sporotrichioides_cDNA_clone_c4c07fs_3',_mRNA_sequen BI188400_c4c07fs.f1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_c4c07fs_3',_mRNA_sequenBI188400_c4c07fs.f1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_c4c07fs_3',_mRNA_sequen 323.2

AFC_ECORC_Fusarium_graminearum_mycelium_cornmeal_solid_culture_Gibberella_zeae_cDNA_clone_Fg05_03g18,_mRNA_seque CD457047_Fg05_03g18_R_Fg05_AAFC_ECORC_Fusarium_graminearum_mycelium_cornmeal_solid_culture_Gibberella_zeae_cDNA_clone_Fg05_03g18,_mRNA_sequeBI188401_c4c07fs.r1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_c4c07fs_5',_mRNA_sequen 323.4

DY989118_GpanSEQ6433_Geomyces_pannorum_pBluescript_(EcoRI-XhoI)_Geomyces_pannorum_cDNA_clone_GpanSEQ6433,_mRNA_sequence 323.8DY588253_DD1_Fusarium_culmorum_Differential_Display_library_Fusarium_culmorum_cDNA,_mRNA_sequence 323.9

324.8_Fusarium_graminearum_mycelium_full-length_cDNA_Gibberella_zeae_cDNA_clone_Fg12_01e22,_mRNA_sequence 325.28,_mRNA_sequence BU063135_Fgr_2_L12_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequencetrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_n3e05fs_3',_mRNA_sequen

DR641601_EST1032226_FvM_Gibberella_moniliformis_cDNA_clone_FVMCC30,_mRNA_sequence325.3

A,_mRNA_sequence BU065054_Fgr_6_I05_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequenceBU062871_Fgr_2_F20_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 325.4

mRNA_sequence DR643256_EST1033881_FvM_Gibberel la_moniliformis_cDNA_clone_FVMCX07,_mRNA_sequence 325.6BU061143_FgrN_5_E04_T7_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 326.3

_06g04,_mRNA_seq BU064587_Fgr_5_E04_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence2,_mRNA_sequence BU064587_Fgr_5_E04_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence

DR645219_EST1035844_FvM_Gibberella_moniliformis_cDNA_clone_FVMDL62,_mRNA_sequence326.4

DR671208_EST1061325_FvO_Gibberella_moniliformis_cDNA_clone_FVOBG27,_mRNA_sequence 326.508_02h06_R_Fg08_AAFC_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_02h06,_mRNA_sequence CD459340_Fg08_02h06_A_Fg08_AAFC_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_02h06,_mRNA_sequence

_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_02h06,_mRNA_sequence 326.6

C_ECORC_Fusarium_graminearum_simple_substrate_Gibberella_zeae_cDNA_clone_Fg09_07o03,_mRNA_sequence 326.7_mRNA_sequence DR648628_EST1038745_FvN_Gibberella_moniliformis_cDNA_clone_FVNAH88,_mRNA_sequence 326.8_mRNA_sequence DR651955_EST1042072_FvN_Gibberella_moni liformis_cDNA_clone_FVNB342,_mRNA_sequence 326.9essed_on_plant_cell_walls_DIFCW_SSH_cDNA_Gibberella_zeae_cDNA_clone_ae07_similar_to_gb:AAO49455.1_|_h646389_mycelial_cDNA_library_Fusarium_oxysporum_f._sp._melonis_cDNA_clone_WVG-14_G11,_mRNA_sequence 327.0

_mRNA_sequence BU060077_Fgr-C_0_K11_T3_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequenceBU060078_Fgr-C_0_K11_T7_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 327.1

C_ECORC_Fusarium_graminearum_simple_substrate_Gibberella_zeae_cDNA_clone_Fg09_05a03,_mRNA_sequence 327.2trichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_b2e06fs_5',_mRNA_sequenmRNA_sequence DR673008_EST1063125_FvO_Gibberella_monil iformis_cDNA_clone_FVOBR87,_mRNA_sequence 327.3

mRNA_sequence DR640169_EST1030794_FvM_Gibberella_moniliformis_cDNA_clone_FVMBU66,_mRNA_sequenceDR656825_EST1046942_FvN_Gibberella_moniliformis_cDNA_clone_FVNBY55,_mRNA_sequence 327.4

trichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_g1c04fs_5',_mRNA_sequen 327.5Scaffold1.49 327.7

',_mRNA_sequen DR678584_EST1068701_FvO_Gibberella_monil iformis_cDNA_clone_FVOCS27,_mRNA_sequence_mRNA_sequence DR649105_EST1039222_FvN_Gibberella_moniliformis_cDNA_clone_FVNAK86,_mRNA_sequence 327.9

agnaporthe_grisea_CS_Uni-Zap_XR_Library_Magnaporthe_grisea_cDNA_clone_mgcs005xO05_5',_mRNA_sequence 328.2_Fusarium_graminearum_mycelium_full-length_cDNA_Gibberella_zeae_cDNA_clone_Fg14_01e08,_mRNA_sequence 328.4

g14_ 2_AAFC_ECORC_Fusarium_graminearum_mycelium_Gibberella_zeae_cDNA_clone_Fg02_04g07,_mRNA_sequence 328.57,_mRNA_sequence BU064858_Fgr_5_P19_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequenceC_ECORC_Fusarium_graminearum_simple_substrate_Gibberella_zeae_cDNA_clone_Fg09_03d05,_mRNA_sequence 328.6

_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_06e01,_mRNA_sequence 328.708_06e01_A_Fg08_AAFC_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_06e01,_mRNA_sequence CD459266_Fg08_06e01_A_Fg08_AAFC_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_06e01,_mRNA_sequence

_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_06e01,_mRNA_sequence 328.8

arium_graminearum_mycelium_grown_on_wheat_heads_Gibberella_zeae_cDNA_clone_Fg04d_01a13,_mRNA_seque 329.0DNA,_mRNA_sequence BU068760_2567_E03_I05ZT5_Mature_peri thecia_Gibberella_zeae_cDNA,_mRNA_sequence 329.2

DR653129_EST1043246_FvN_Gibberella_moniliformis_cDNA_clone_FVNBA79,_mRNA_sequence 329.3DR644705_EST1035330_FvM_Gibberella_moniliformis_cDNA_clone_FVMDF29,_mRNA_sequence 329.7

_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_03c04,_mRNA_sequence 329.8458146_Fg08_10d09_A_Fg08_AAFC_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_10d09,_mRNA_sequence BI749718_Fg02_04b06_R_Fg02_AAFC_ECORC_Fusarium_graminearum_mycelium_Gibberella_zeae_cDNA_clone_Fg02_04b06,_mRNA_sequence

2_AAFC_ECORC_Fusarium_graminearum_mycelium_Gibberella_zeae_cDNA_clone_Fg02_04b06,_mRNA_sequence 329.9

mRNA_sequence DR637099_EST1027724_FvM_Gibberella_moniliformis_cDNA_clone_FVMAU03,_mRNA_sequencemRNA_sequence DR637099_EST1027724_FvM_Gibberella_moniliformis_cDNA_clone_FVMAU03,_mRNA_sequence 330.2

A,_mRNA_sequence BU064184_Fgr_4_D06_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 330.3BU065549_Fgr_7_N22_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 330.5

DR611298_EST1001426_FvG_Gibberella_moniliformis_cDNA_clone_FVGDX83,_mRNA_sequence 331.1Fusarium_graminearum_mycelium_cornmeal_solid_culture_Gibberella_zeae_cDNA_clone_Fg05_01l01,_mRNA_seque 331.2RC_Fusarium_graminearum_mycelium_partial_cDNA_Gibberella_zeae_cDNA_clone_Fg13_01e18,_mRNA_sequence 331.3

5_Fgr-S4_3_G06_T7_F._graminearum_infected_wheat_subtraction_library_Gibberella_zeae_cDNA,_mRNA_sequence 331.7_mRNA_sequence DR662580_EST1052697_FvN_Gibberella_moniliformis_cDNA_clone_FVND901,_mRNA_sequence 332.6

CN813160_Fg06_04o14_R_Fg06_AAFC_ECORC_Fusarium_graminearum_perithecia_Gibberella_zeae_cDNA_clone_Fg06_04o14,_mRNA_sequence CN813160_Fg06_04o14_R_Fg06_AAFC_ECORC_Fusarium_graminearum_perithecia_Gibberella_zeae_cDNA_clone_Fg06_04o14,_mRNA_sequence6_AAFC_ECORC_Fusarium_graminearum_perithecia_Gibberella_zeae_cDNA_clone_Fg06_04o14,_mRNA_sequence 333.9

Scaffold1.51 334.3

Scaffold1.52 334.6_mRNA_sequence DR654449_EST1044566_FvN_Gibberella_moniliformis_cDNA_clone_FVNBJ32,_mRNA_sequence 334.7

_Fusarium_graminearum_mycelium_full-length_cDNA_Gibberella_zeae_cDNA_clone_Fg14_03g23,_mRNA_sequence 335.13,_mRNA_sequence BU064159_Fgr_4_C04_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 335.2

DR630057_EST1020185_FvI_Gibberella_moniliformis_cDNA_clone_FVIDL67,_mRNA_sequence 336.7

DY763079_DD10a_Fusarium_graminearum_Differential_Display_library_Gibberella_zeae_cDNA,_mRNA_sequence 337.7BU065069_Fgr_6_I20_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 337.8

_ECORC_Fusarium_graminearum_simple_substrate_Gibberella_zeae_cDNA_clone_Fg09_01m18,_mRNA_sequence 337.9

DR647838_EST1037955_FvN_Gibberella_moniliformis_cDNA_clone_FVNAC75,_mRNA_sequence 339.4

C_ECORC_Fusarium_graminearum_simple_substrate_Gibberella_zeae_cDNA_clone_Fg09_06a22,_mRNA_sequence 341.0C_ECORC_Fusarium_graminearum_simple_substrate_Gibberella_zeae_cDNA_clone_Fg09_06a22,_mRNA_sequence 341.145044_conidial_cDNA_library_Fusarium_oxysporum_f._sp._melonis_cDNA_clone_WCMC-15_C05,_mRNA_sequence 341.5

3',_mRNA_sequen DR667788_EST1057905_FvO_Gibberella_moniliformis_cDNA_clone_FVOAT37,_mRNA_sequence 341.60_Fg09_05c18_A_Fg09_AAFC_ECORC_Fusarium_graminearum_simple_substrate_Gibberella_zeae_cDNA_clone_Fg09_05c18,_mRNA_sequence CD460870_Fg09_05c18_A_Fg09_AAFC_ECORC_Fusarium_graminearum_simple_substrate_Gibberella_zeae_cDNA_clone_Fg09_05c18,_mRNA_sequence 341.7Fg09_ pressed_on_plant_cell_walls_DIFCW_SSH_cDNA_Gibberella_zeae_cDNA_clone_3E07_similar_to_Protein_expressed 341.807_-_differentiall y_expressed_on_plant_cell_walls_DIFCW_SSH_cDNA_Gibberella_zeae_cDNA_clone_3E07_similar_to_Protein_expressed CN811281_Fg13_02c05_T3_Fg13_AAFC_ECORC_Fusarium_graminearum_mycelium_partial_cDNA_Gibberella_zeae_cDNA_clone_Fg13_02c05,_mRNA_sequence 341.9T3_Fg13_AAFC_ECORC_Fusarium_graminearum_mycelium_partial_cDNA_Gibberella_zeae_cDNA_clone_Fg13_02c05,_mRNA_sequence CN811281_Fg13_02c05_T3_Fg13_AAFC_ECORC_Fusarium_graminearum_mycelium_partial_cDNA_Gibberella_zeae_cDNA_clone_Fg13_02c05,_mRNA_sequence 342.0

03,_mRNA_seque DR679378_EST1069495_FvO_Gibberella_moniliformis_cDNA_clone_FVOCX54,_mRNA_sequenceDR628236_EST1018364_FvI_Gibberella_moniliformis_cDNA_clone_FVICQ58,_mRNA_sequence 342.5

5,_mRNA_sequence DR631079_EST1021207_FvI_Gibberella_monil iformis_cDNA_clone_FVIDZ69,_mRNA_sequenceDR597381_EST987509_FvF_Gibberella_moniliformis_cDNA_clone_FVFIY60,_mRNA_sequence 342.6

BU066227_Fgr_9_L03_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 343.1ST_9f09_-_differentially_expressed_on_plant_cell_walls_DIFCW_SSH_cDNA_Gibberella_zeae_cDNA_clone_9f09_similar_to_sp:P06785_|_Thymi CV828207_EST_9f09_-_differentially_expressed_on_plant_cell_wal ls_DIFCW_SSH_cDNA_Gibberella_zeae_cDNA_clone_9f09_similar_to_sp:P06785_|_Thymi

DR595949_EST986077_FvF_Gibberella_monil iformis_cDNA_clone_FVFIH49,_mRNA_sequence 343.2

_mRNA_sequence DR656774_EST1046891_FvN_Gibberella_moniliformis_cDNA_clone_FVNBY19,_mRNA_sequenceDR678401_EST1068518_FvO_Gibberella_moniliformis_cDNA_clone_FVOCR17,_mRNA_sequence 343.5

DR636807_EST1027432_FvM_Gibberella_monil iformis_cDNA_clone_FVMAS07,_mRNA_sequence 343.6CN814307_Fg06_05d08_R_Fg06_AAFC_ECORC_Fusarium_graminearum_perithecia_Gibberella_zeae_cDNA_clone_Fg06_05d08,_mRNA_sequence CN814307_Fg06_05d08_R_Fg06_AAFC_ECORC_Fusarium_graminearum_perithecia_Gibberella_zeae_cDNA_clone_Fg06_05d08,_mRNA_sequence 343.7

DR618548_EST1008676_FvH_Gibberella_moniliformis_cDNA_clone_FVHDG59,_mRNA_sequence 343.8RC_Fusarium_graminearum_mycelium_partial_cDNA_Gibberella_zeae_cDNA_clone_Fg13_02a04,_mRNA_sequence 343.9_thermophile_pBluescript_(EcoRI-XhoI)_Corynascus_heterothal licus_cDNA_clone_StheSEQ14438,_mRNA_sequence 344.4

_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_03h06,_mRNA_sequence 344.6g08_03h06_A_Fg08_AAFC_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_03h06,_mRNA_sequence CD459123_Fg08_05c11_A_Fg08_AAFC_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_05c11,_mRNA_sequence 344.7

BU066896_1612_D10_H20ZT_Mature_peri thecia_Gibberella_zeae_cDNA,_mRNA_sequence 344.8_p1g trichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_p1g07fs_5',_mRNA_sequen 344.992139_m1d07fs.r1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_m1d07fs_5',_mRNA_sequen BI750168_Fg02_10c05_R_Fg02_AAFC_ECORC_Fusarium_graminearum_mycelium_Gibberella_zeae_cDNA_clone_Fg02_10c05,_mRNA_sequence

DR674938_EST1065055_FvO_Gibberella_moniliformis_cDNA_clone_FVOC471,_mRNA_sequence CN811439_Fg14_02l02_C_Fg14_AAFC_ECORC_Fusarium_graminearum_mycelium_full-length_cDNA_Gibberella_zeae_cDNA_clone_Fg14_02l02,_mRNA_sequence 345.2

07e02_A_Fg09_AAFC_ECORC_Fusarium_graminearum_simple_substrate_Gibberella_zeae_cDNA_clone_Fg09_07e02,_mRNA_sequence EB530859_Fg13_03g21_T3_Fg13_AAFC_ECORC_Fusarium_graminearum_mycelium_partial_cDNA_Gibberella_zeae_cDNA_clone_Fg13_03g21,_mRNA_sequence 345.3CN813014_Fg06_01p16_R_Fg06_AAFC_ECORC_Fusarium_graminearum_perithecia_Gibberella_zeae_cDNA_clone_Fg06_01p16,_mRNA_sequence CN813014_Fg06_01p16_R_Fg06_AAFC_ECORC_Fusarium_graminearum_perithecia_Gibberella_zeae_cDNA_clone_Fg06_01p16,_mRNA_sequence 345.5CN813014_Fg06_01p16_R_Fg06_AAFC_ECORC_Fusarium_graminearum_perithecia_Gibberella_zeae_cDNA_clone_Fg06_01p16,_mRNA_sequence CN813014_Fg06_01p16_R_Fg06_AAFC_ECORC_Fusarium_graminearum_perithecia_Gibberella_zeae_cDNA_clone_Fg06_01p16,_mRNA_sequence

BU060593_Fgr-C_1_F13_T7_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 345.6

BU060593_Fgr-C_1_F13_T7_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence BU060061_Fgr-C_0_K03_T3_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequenceBU060061_Fgr-C_0_K03_T3_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 345.7

A_sequence DR611358_EST1001486_FvG_Gibberella_monil iformis_cDNA_clone_FVGDY56,_mRNA_sequence 346.5

RNA_sequence DR597223_EST987351_FvF_Gibberella_moniliformis_cDNA_clone_FVFIW78,_mRNA_sequence 346.9

ORC_Fusarium_graminearum_simple_substrate_Gibberella_zeae_cDNA_clone_Fg09_04a21,_mRNA_sequence 347.5_Fg09_04a21_A_Fg09_AAFC_ECORC_Fusarium_graminearum_simple_substrate_Gibberella_zeae_cDNA_clone_Fg09_04a21,_mRNA_sequence CD461322_Fg09_04a21_A_Fg09_AAFC_ECORC_Fusarium_graminearum_simple_substrate_Gibberella_zeae_cDNA_clone_Fg09_04a21,_mRNA_sequence 347.6

NA_sequence BU059951_Fgr-C_0_H17_T7_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 348.8NA_sequence BU059950_Fgr-C_0_H17_T3_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence

Scaffold1.53 BU066194_Fgr_9_J18_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequenceBU066194_Fgr_9_J18_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence

348.9

A_sequence DR663681_EST1053798_FvO_Gibberella_moniliformis_cDNA_clone_FVOA077,_mRNA_sequenceides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_p3b01fs_5',_mRNA_sequen 349.0

BU060184_Fgr-C_0_M20_T7_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 349.2RNA_sequence BU064228_Fgr_4_F02_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence

BU064228_Fgr_4_F02_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 349.3

a_zeae_cDNA,_mRNA_sequence BQ110405_VD0109H01_VD01_Vertici llium_dahliae_cDNA,_mRNA_sequenceides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_b4a03fs_5',_mRNA_sequen

A_sequence BU059415_Fgr-C_02_G23_T3_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence349.4

4a ed_on_plant_cell_walls_DIFCW_SSH_cDNA_Gibberel la_zeae_cDNA_clone_3g05_similar_to_Protein_expressed 349.5RNA_sequence BU065252_Fgr_7_A23_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence

BU065252_Fgr_7_A23_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 349.6

RC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_12g12,_mRNA_sequence 349.7NA_sequence BU060802_Fgr-C_1_J22_T3_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence

BU060802_Fgr-C_1_J22_T3_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 349.8

BU060802_Fgr-C_1_J22_T3_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 349.9_mRNA_sequence BU067050_1614_E02_I04Z5T_Mature_peri thecia_Gibberella_zeae_cDNA,_mRNA_sequence

BU060121_Fgr-C_0_L09_T7_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 350.5

RNA_sequence BU065906_Fgr_8_N06_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 350.6BU068727_2565_H11_P21ZT5_Mature_peri thecia_Gibberella_zeae_cDNA,_mRNA_sequence 350.9

mRNA_sequence BU068307_2556_H01_O02ZT5_Mature_peri thecia_Gibberella_zeae_cDNA,_mRNA_sequencemRNA_sequence BU066468_1606_G11_M22ZS5_Mature_peri thecia_Gibberella_zeae_cDNA,_mRNA_sequence 351.1

RNA_sequence BU065145_Fgr_6_M06_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 351.2RNA_sequence BU065145_Fgr_6_M06_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequenceRNA_sequence BU065145_Fgr_6_M06_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence4839_mycelium_Lambda_Zap_Library_L02_Hypocrea_lixii_cDNA_clone_L02T34P053R04891,_mRNA_sequence

351.3

BU066871_1612_B05_D10ZT_Mature_peri thecia_Gibberella_zeae_cDNA,_mRNA_sequence 351.6BU067325_1619_D10_H19ZT5_Mature_peri thecia_Gibberella_zeae_cDNA,_mRNA_sequence 351.7

um_graminearum_mycelium_trichothecene_production_Gibberella_zeae_cDNA_clone_Fg03_05e04,_mRNA_seq 352.9DR597233_EST987361_FvF_Gibberella_moniliformis_cDNA_clone_FVFIW89,_mRNA_sequence 353.0

A_sequence DR638815_EST1029440_FvM_Gibberella_moniliformis_cDNA_clone_FVMBD58,_mRNA_sequenceDR638815_EST1029440_FvM_Gibberella_moniliformis_cDNA_clone_FVMBD58,_mRNA_sequence 353.5

Y763133_DD11i_Fusarium_graminearum_Differential_Display_library_Gibberella_zeae_cDNA,_mRNA_sequence 353.7aporthe_grisea_NS_Uni-Zap_XR_Library_Magnaporthe_grisea_cDNA_clone_mgns015xJ04_5',_mRNA_sequence 354.1NA_sequence BU060276_Fgr-C_0_O22_T3_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence

BU060277_Fgr-C_0_O22_T7_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 354.2

oides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_k2h05fs_5',_mRNA_sequen 354.3NA_sequence DR671478_EST1061595_FvO_Gibberella_moniliformis_cDNA_clone_FVOBI01,_mRNA_sequence

DR665963_EST1056080_FvO_Gibberella_monil iformis_cDNA_clone_FVOAH38,_mRNA_sequence 354.7

RNA_sequence BU063495_Fgr_3_D02_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequenceRNA_sequence BU063496_Fgr_3_D02_T7_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence

BU063496_Fgr_3_D02_T7_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence354.9

um_graminearum_mycelium_trichothecene_production_Gibberella_zeae_cDNA_clone_Fg03_08c01,_mRNA_seq 355.0AFC_ECORC_Fusarium_graminearum_mycelium_trichothecene_production_Gibberella_zeae_cDNA_clone_Fg03_08c01,_mRNA_seq CD456084_Fg03_08c01_R_Fg03_AAFC_ECORC_Fusarium_graminearum_mycelium_trichothecene_production_Gibberella_zeae_cDNA_clone_Fg03_08c01,_mRNA_seq 355.1

RNA_sequence BU064319_Fgr_4_I21_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequenceRNA_sequence BU064319_Fgr_4_I21_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence

BU064319_Fgr_4_I21_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence355.3

RNA_sequence BU063299_Fgr_2_O22_T7_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 355.4Scaffold1.54 355.5

RC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_11g10,_mRNA_sequence 356.4mRNA_sequence BU067470_1625_D02_G03ZT5_Mature_peri thecia_Gibberella_zeae_cDNA,_mRNA_sequence 356.5A_sequence DR610254_EST1000382_FvG_Gibberella_moniliformis_cDNA_clone_FVGDH55,_mRNA_sequence 356.6mRNA_sequen BU062576_Fgr_1_P15_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence

DR623439_EST1013567_FvI_Gibberella_moniliformis_cDNA_clone_FVIAQ58,_mRNA_sequence 356.7

RNA_sequence BU062577_Fgr_1_P15_T7_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 356.8FC_ECORC_Fusarium_graminearum_perithecia_Gibberella_zeae_cDNA_clone_Fg06_02g16,_mRNA_sequenceFC_ECORC_Fusarium_graminearum_perithecia_Gibberella_zeae_cDNA_clone_Fg06_03n06,_mRNA_sequence 357.0

BU059472_Fgr-C_02_J09_T3_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 357.2BU059472_Fgr-C_02_J09_T3_Carbon-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence 357.3DR680041_EST1070158_FvO_Gibberella_moniliformis_cDNA_clone_FVOD203,_mRNA_sequence 357.5

EB511559_Fg_Ta07b_04e09_R_Fg_Ta07b_AAFC_ECORC_Fusarium_graminearum_inculated_wheat_heads_Gibberella_zeae_cDNA_clone_Fg_Ta07b_04e09,_mRNA_seq 357.9832_l3a02fs.f1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_l3a02 fs_3',_mRNA_sequen BI191832_l3a02fs.f1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_l3a02fs_3',_mRNA_sequen 358.0

BI191833_l3a02fs.r1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_l3a02fs_5',_mRNA_sequen 358.1DR659699_EST1049816_FvN_Gibberella_moniliformis_cDNA_clone_FVNCH05,_mRNA_sequence DR659699_EST1049816_FvN_Gibberella_moniliformis_cDNA_clone_FVNCH05,_mRNA_sequence 358.9

2e03fs.r1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_m2e03fs_5',_mRNA_sequen BI192265_m2e03fs.r1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_m2e03fs_5',_mRNA_sequenBI192265_m2e03fs.r1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_m2e03fs_5',_mRNA_sequen 359.2

EB530591_Fg13_02e24_T3_Fg13_AAFC_ECORC_Fusarium_graminearum_mycelium_partial_cDNA_Gibberella_zeae_cDNA_clone_Fg13_02e24,_mRNA_sequence DR635985_EST1026610_FvM_Gibberella_moniliformis_cDNA_clone_FVMAM59,_mRNA_sequence 359.3_01a16_T3_Fg13_AAFC_ECORC_Fusarium_graminearum_mycelium_partial_cDNA_Gibberella_zeae_cDNA_clone_Fg13_01a16,_mRNA_sequence BI191891_l3d01fs.f1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_l3d01fs_3',_mRNA_sequen 359.4

DR677377_EST1067494_FvO_Gibberella_monil iformis_cDNA_clone_FVOCK66,_mRNA_sequence 359.6CD458152_Fg08_11k17_A_Fg08_AAFC_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_11k17,_mRNA_sequence 359.7

g08_11k17_A_Fg08_AAFC_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_11k17,_mRNA_sequence CD458641_Fg08_12p19_A_Fg08_AAFC_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_12p19,_mRNA_sequenceCD458641_Fg08_12p19_A_Fg08_AAFC_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_12p19,_mRNA_sequence 359.8

08_12p19_A_Fg08_AAFC_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_12p19,_mRNA_sequence CD458641_Fg08_12p19_A_Fg08_AAFC_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_12p19,_mRNA_sequence 359.9

1 1

6O11

5D232D13

5B115J13

4J46F1

2I11

5O193L10

4L13

6F33G

226e_16

3D202H23

3A6

4L144J5

1P215I21

6A244H14

4P4

6J143O

145e_14

6N95e_11

3D202M

116O

3

5G20

2I11e_22

1H231P21

6L146e_11

6H11

1N246H23

2O10

5H21

3L10

6F3

Fg09_0

a03fs.f

FC_EC

CN8

DR665464_EST1055581_FvO_Gibberella_moniliformis_cDNA_clone_FVOAE20,_BU063136_Fgr_2_L12_T7_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence CN812550_Fg12_01e22_T3_Fg12_AAFC_ECORC_CN812507_Fg12_01c08_T7_Fg12_AAFC_ECORC_Fusarium_graminearum_mycelium_full-length_cDNA_Gibberella_zeae_cDNA_clone_Fg12_01c08

DR641601_EST1032226_FvM_Gibberel la_moniliformis_cDNA_clone_FVMCC30,_mRNA_sequence BI200520_n3e05fs.f1_Fusarium_sporot

BU065054_Fgr_6_I05_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA

DR643256_EST1033881_FvM_Gibberella_moniliformis_cDNA_clone_FVMCX07,_m

CD456021_Fg03_06g04_R_Fg03_AAFC_ECORC_Fusarium_graminearum_mycelium_trichothecene_production_Gibberella_zeae_cDNA_clone_Fg03_DR645219_EST1035844_FvM_Gibberella_moniliformis_cDNA_clone_FVMDL62

02h06_R_Fg08_AAFC_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_02h06,_mRNA_sequence CD459340_Fg08_02h06_A_Fg08_AAFC_CD459340_Fg08_02h06_A_Fg08_AAFC_

BU066310_Fgr_9_O17_T3_Nitrogen-sta rved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence CD459961_Fg09_07o03_A_Fg09_AAFCBU065913_Fgr_8_N14_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_BU063661_Fgr_3_G14_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_

DR678016_EST1068133_FvO_Gibberel la_moniliformis_cDNA_clone_FVOCO72,_mRNA_sequence CV828278_EST_ae07_-_differentially_exprBW646389_BW6

BI750230_Fg02_06b12_R_Fg02_AAFC_ECORC_Fusarium_graminearum_mycelium_Gibberella_zeae_cDNA_clone_Fg02_06b12,_

BU061383_Fgr_0_I11_T7_Nitrogen-sta rved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence CD461036_Fg09_05a03_A_Fg09_AAFCDR673008_EST1063125_FvO_Gibberella_moniliformis_cDNA_clone_FVOBR87,_mRNA_sequence BI187803_b2e06fs.r1_Fusarium_sporot

DR673008_EST1063125_FvO_Gibberel la_moniliformis_cDNA_clone_FVOBR87,_DR649598_EST1039715_FvN_Gibberella_moniliformis_cDNA_clone_FVNAO20,_m

DR656825_EST1046942_FvN_Gibberella_moniliformis_cDNA_clone_FVNBY55,_mRNA_sequence BI189725_g1c04fs.r1_Fusarium_sporot

BI190730_i4c03fs.r1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_i4c03fs_5DR649105_EST1039222_FvN_Gibberella_moniliformis_cDNA_clone_FVNAK86,_

Scaffold1.50 BM866260_mgcs005xO05f.b_MEB530988_Fg14_01e08_C_Fg14_AAFC_ECORC_

_01e08_C_Fg14_AAFC_ECORC_Fusarium_graminearum_mycelium_full-length_cDNA_Gibberella_zeae_cDNA_clone_Fg14_01e08,_mRNA_sequence BI749851_Fg02_04g07_R_Fg02BI749851_Fg02_04g07_R_Fg02_AAFC_ECORC_Fusarium_graminearum_mycelium_Gibberella_zeae_cDNA_clone_Fg02_04g07

CD459394_Fg09_03d05_A_Fg09_AAFCCD459266_Fg08_06e01_A_Fg08_AAFC_

06e01_A_Fg08_AAFC_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_06e01,_mRNA_sequence CD459266_Fg08_06e01_A_Fg08_AAFC_CD459266_Fg08_06e01_A_Fg08_AAFC_

CD457899_Fg04d_01a13_A_Fg04_AAFC_ECORC_FusaBU068760_2567_E03_I05ZT5_Mature_perithecia_Gibberella_zeae_cD

CD458802_Fg08_03c04_A_Fg08_AAFC_46_Fg08_10d09_A_Fg08_AAFC_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_10d09,_mRNA_sequence BI749718_Fg02_04b06_R_Fg02

DR629874_EST1020002_FvI_Gibberella_moniliformis_cDNA_clone_FVIDJ04,_mRNA_sequence BI749718_Fg02_04b06_R_Fg02DR678151_EST1068268_FvO_Gibberella_moniliformis_cDNA_clone_FVOCP55,_mDR637100_EST1027725_FvM_Gibberella_moniliformis_cDNA_clone_FVMAU03,_m

BU064184_Fgr_4_D06_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA

CN812136_Fg05_01l01_R_Fg05_AAFC_ECORC_FCN811557_Fg13_01e18_T3_Fg13_AAFC_ECO

DV998695DR662580_EST1052697_FvN_Gibberella_moniliformis_cDNA_clone_FVND901,_

CN813160_Fg06_04o14_R_Fg06_AAFC_ECORC_Fusarium_graminearum_perithecia_Gibberella_zeae_cDNA_clone_Fg06_04o14,_mRNA_sequence CN813160_Fg06_04o14_R_Fg06CN813160_Fg06_04o14_R_Fg06

DR654449_EST1044566_FvN_Gibberella_moniliformis_cDNA_clone_FVNBJ32,_

CN811361_Fg14_03g23_A_Fg14_AAFC_ECORC_CN811361_Fg14_03g23_A_Fg14_AAFC_ECORC_Fusarium_graminearum_mycelium_full-length_cDNA_Gibberella_zeae_cDNA_clone_Fg14_03g23

BU065069_Fgr_6_I20_T3_Nitrogen-starved_mycelia_Gibberella_zeae_cDNA,_mRNA_sequence CD460796_Fg09_01m18_A_Fg09_AAFC

CD460352_Fg09_06a22_A_Fg09_AAFCBU068187_2555_C01_E01ZT5_Mature_perithecia_Gibberella_zeae_cDNA,_mRNA_sequence CD460352_Fg09_06a22_A_Fg09_AAFC

BI191452_k3g02fs.r1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_k3g02fs_5',_mRNA_sequen BW645044_BW64BI191451_k3g02fs.f1_Fusarium_sporotrichio ides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_k3g02fs_3

09_05c18_A_Fg09_AAFC_ECORC_Fusarium_graminearum_simple_substrate_Gibberella_zeae_cDNA_clone_Fg09_05c18,_mRNA_sequence CD460870_Fg09_05c18_A_Fg09_AAFC_05c18_A_Fg09_AAFC_ECORC_Fusarium_graminearum_simple_substrate_Gibberella_zeae_cDNA_clone_Fg09_05c18,_mRNA_sequence CV827702_EST_3E07_-_differentially_exp_differentiall y_expressed_on_plant_cell_walls_DIFCW_SSH_cDNA_Gibberella_zeae_cDNA_clone_3E07_similar_to_Protein_expressed CN811281_Fg13_02c05_T3_Fg13_AAFC_ECOg13_AAFC_ECORC_Fusarium_graminearum_mycelium_partial_cDNA_Gibberella_zeae_cDNA_clone_Fg13_02c05,_mRNA_sequence CN811281_Fg13_02c05_T3_Fg13_AAFC_ECO

CD457952_Fg04d_01k03_A_Fg04_AAFC_ECORC_Fusarium_graminearum_mycelium_grown_on_wheat_heads_Gibberella_zeae_cDNA_clone_Fg04d_01k0

DR609996_EST1000124_FvG_Gibberella_moniliformis_cDNA_clone_FVGDD95

f09_-_differentially_expressed_on_plant_cell_walls_DIFCW_SSH_cDNA_Gibberella_zeae_cDNA_clone_9f09_similar_to_sp:P06785_|_Thymi CV828207_EST_9f09_-_differentially_exp

DR656774_EST1046891_FvN_Gibberella_moniliformis_cDNA_clone_FVNBY19,_

CN814307_Fg06_05d08_R_Fg06_AAFC_ECORC_Fusarium_graminearum_perithecia_Gibberella_zeae_cDNA_clone_Fg06_05d08,_mRNA_sequence CN814307_Fg06_05d08_R_Fg06

DR595310_EST985438_FvF_Gibberella_moniliformis_cDNA_clone_FVFIA58,_mRNA_sequence EB530531_Fg13_02a04_T3_Fg13_AAFC_ECOEB054916_StheSEQ14438_Sporotri chum_CD458849_Fg08_03h06_A_Fg08_AAFC_

03h06_A_Fg08_AAFC_ECORC_Fusarium_graminearum_complex_substrate_Gibberella_zeae_cDNA_clone_Fg08_03h06,_mRNA_sequence CD459123_Fg08_05c11_A_Fg08_AAFC_

g07fs.f1_Fusarium_sporotrichioides_Tri_10_overexpressed_cDNA_library_Fusarium_sporotrichioides_cDNA_clone_p1g07fs_3',_mRNA_sequen BI201265_p1g07fs.r1_Fusarium_sporot39 m1d07fs r1 Fusarium sporotrichioides Tri 10 overexpressed cDNA library Fusarium sporotrichioides cDNA clone m1d07fs 5' mRNA sequen BI750168 Fg02 10c05 R Fg0

Sequence Alignment

Color Key

HindIII restriction site

HindIII restriction site

BMC Genomics 2007, 8:262 http://www.biomedcentral.com/1471-2164/8/262

gies to reduce yield loss to SDS [4] may also be developed.For example, because F. virguliforme is a member of theFusarium plant disease complex which contains manyrelated pathogens of all the worlds major crops [5,6],development of genome based fungicides and seed treat-ments for one may have broad utility. Since F. virguliformeis a hemibiotrophic plant pathogen with a wide hostrange [1,5], tests against many model plant species toidentify new resistance genes are underway. Further, sincethe perfect stage is not known and all extant strains appearto be of a single mating type [7], these maps represent asignificant genetic tool. Finally, the BAC libraries presentopportunities for functional genomics, since the vector isa binary T-DNA capable of both plant and fungal transfor-mation mediated by Agrobacterium strains [8,9].

ConclusionThe main power of this method lay in its ability to aligndiverged sequences. Because the program uses relation-ships between 15 bp sequences, the reference genome canexhibit significant change from the probe sequences, yetstill effectively group fragments of DNA. This work isunique in that discontinuous sequences were used for theanalysis, information not readily apparent such as matchdirection are presented and the JAVA programs used arefreely available. The 94 maps represent an excellentresource for continuing research on Fusarium virguliforme.

AbbreviationsBAC, bacterial artificial chromosome; BES, bacterial artifi-cial chromosome end sequence; bp, base pair; SDS, soy-bean sudden death syndrome; SNP, single nucleotidepolymorphism.

Authors' contributionsJS conceived of the study; SA and DL provided criticalreview, interpretation of results and funding (DL), LB per-formed geneic annotation of the F. graminearumsequences. All authors read and agreed to the final manu-script.

Additional material

AcknowledgementsThanks are due to Dr Javed Iqbal, Dr. Chris Town, Dr. K Meksem, Hiro Ishihara, H. Koo, J. Yuan, AJ Afzal and Rubina Ahsan for BAC libraries and BES data. The research was funded in part by grants from United Soybean Board to MJI and DAL. Any opinions and findings were of authors and USB is not responsible for the content. Physical map development was based upon work supported by the National Science Foundation under Grant No. 9872635. Any opinions, findings, and conclusions or recommendations expressed in this material are those of the author(s) and do not necessarily reflect the views of the National Science Foundation. The continued sup-port of SIUC, College of Agriculture and Office of the Vice Chancellor for Research to DAL is appreciated.

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12. Shultz J, Meksem K, Shetty J, Town CD, Koo H, Potter J, WakefieldK, Zhang H, Wu C, Lightfoot DA: End sequencing of BACs com-prising a provisional tiling path from a fingerprint physicalmap of soybean (Glycine max) cultivar Forrest. Genbank.CG812653 to CG826126 (13,473 sequences). 2003.

13. Budiman MA, Mao L, Wood TC, Wing RA: A deep-coveragetomato BAC library and prospects toward development ofan STC framework for genome sequencing. Genome Res 2000,10(1):129-136.

14. Guldener U, Mannhaupt G, Munsterkotter M, Haase D, OesterheldM, Stumpflen V, Mewes HW, Adam G: FGDB: a comprehensivefungal genome resource on the plant pathogen Fusariumgraminearum. Nucleic Acids Res 2006, 34(Databaseissue):D456-8.

15. DOE: Joint Genome Institute . 2007 [http://www.jgi.doe.gov/].

Additional file 1A self-extracting file containing 94 maps generated by short sequence comparisons between Fusarium virguliforme and scaffold sequences from Fusarium graminearum. Each map is viewable using the Micro-soft™ Picture and Fax Viewer program. The extracted files include a searchable Microsoft™ Excel file.Click here for file[http://www.biomedcentral.com/content/supplementary/1471-2164-8-262-S1.exe]

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