Patrick X. Zhao, Ph. D.

The Zhao Bioinformatics Lab

pzhao@noble.org

Mapping Affymetrix Medicago GeneChip Probe sets to IMGAG 3.0

Genes

• About Affymetrix Medicago GeneChip

• Mapping Approach

• Bioinformatics & Data Resources for Medicago IMGAG Release V3

Agenda

Affymetrix GeneChip Probes

5’ UTR EXON-I EXON-II EXON-III 3’ UTR

mRNA

Probeset: 11 Probes

Target Sequence

25-mer

1 255 10 15 20

1 255 10 15 20

Perfect match - PM

Mismatch - MM

• id_at:Designates probe sets that uniquely recognize target transcripts

• id_a_at:Designates probe sets that recognize alternative transcripts from the

same gene.• id_s_at:

Designates probe sets with common probes among multiple transcripts from different genes.

• id_x_at: Designates probe sets where it was not possible to select either a

unique probe set or a probe set with identical probes among multiple transcripts. Rules for cross-hybridization were dropped in order to design the _x probe sets. These probe sets share some probes identically with two or more sequences and, therefore, these probe sets may cross-hybridize in an unpredictable manner.

GeneChip® Expression Analysis Data Analysis Fundamentals.

Probeset Types

About Medicago GeneChipType Num of

probe setsPercent in the Mtr. set

Notes

Unique probe sets: e.g. Mtr.10097.1.S1_at

44182 86.80 Unique to one gene

Alternative (_a_), e.g.: Mtr.10267.1.S1_a_at

116 2.28 Alternative probe sets to one gene

Shared (_s_), e.g. Mtr.10146.1.S1_s_at

4793 9.42 Common to multiple genes

Others (_x_), e.g.:Mtr.10093.1.S1_x_at

1809 3.55 Other probe sets with complicated mapping

Total 50900 100

Statistics on Original Medicago GeneChip Probe-sets vs. Gene Index V8 Mapping

Num of ESTs

Matching Probeset

Percent

6315 0 17.12

29038 1 78.74

1525 >=2 4.14

36878 100

• IMGAG V3 gene sequences were matched to corresponding Affymetrix probe sets using a position-weighted scoring index in which mismatches near the middle of a probe were most heavily penalized as follows:

A perfect match for a probe set yields a score of 45

• Matches were declared when at least 8 of 11 probe sets had scores of 43 or higher

Mapping Approach

1 255 10 15 20

[1,1,1,1,1,2,2,2,2,2,3,3,3,3,3,2,2,2,2,2,1,1,1,1,1]

Originated from Affymetrix, Inc.

Num of EST

Matching probe-set

Percent

3304 0 8.96

29535 1 80.09

4039 >=2 10.95

36878 1000

10

20

30

40

50

60

70

80

90

0 probset 1 probeset 2 probesets

OriginOurs

Overlapping mapping between our Probesets vs. Unigene mapping and the Affy original Probesets vs. Unigene mapping. 37872 ∩ 32108=32106. Our method covered 32106/32108=99.9993% of the Affymetrix original mapping.

Statistics on Our Probesets vs. Gene Index V8 Mapping Results

Statistics on Our IMGAG V3 vs. Probesets Mapping Results

Num of cDNA Matching probe_set Percent

29755 0 55.70

16384 1 30.67

7284 >=2 13.63

53423 Total 100

Item Num of probesets

Matched To Percent

1 4860 None 9.552 19709 Unigene only 38.723 24348 19949 Unigene and

unique IMGAGv3

39.19 47.83

4399 Unigene and multiple IMGAGv3

8.64+

4 1983 1698 Unique IMGAGv3 only

3.34 3.90

285 Multiple IMGAGv3 only

0.56++

50900 Total 100

EST 38.72

(47.83)IMGAG

3.90

9.55

Probesets Map to IMGAG V3 and/or Gene Index V9

Medicago Data and Bioinformatics Resources

• http://bioinfo3.noble.org/medicago

AcknowledgementZhao LabXinbin DaiRakesh KaundalHaiquan LiJun LiZhaohong ZhuangJoshua Smith

Collaborators:Michael K. UdvardiRick A. DixonKiran K. MysoreRujin ChenChris Town (JCVI)Greg D. May (NCGR)… …