1. Genome-Wide Analysis of Transposon Insertion Polymorphisms (TIPs) Reveals Intraspecific Variation in Cultivated Rice

2. Sixty Years Ago: A high degree of intraspecific variation in maize caused by “Controlling Element” The TE insertion in the Pigment Gene affect the Synthesis of the Purple Pigment. Feschotte et al, Nat. Rev. Genet. 3 (5):329-341 (2002)

3. 1.Case studies on TE-Induced variation 2.Abundance of TE insertion polymorphisms in rice  Affect Individual Genes  Affect Genome Size 3.Retrotransposon polymorphisms in evolutionary analysis 4.Conclusions Natural Genetic Variation caused by Transposable Elements in cultivated Rice

4. Wheat Xylanase Inhibitor Protein I (XIP-I) Binding to xylanases is necessary for inhibition activity May function in plant defense against secreted fungal pathogen xylanases XIP-I Xylanase Payan et al, J. Biol. Chem. 279 (34):36029-36037 (2004)

5. Tokunaga et al 2007 Goesaert et al 2005 Durand et al 2005 Park et al 2002 The Rice Ortholog of Wheat XIP-I

6. The XIP-I gene split by a LTR retrotransposon in Japonica

7. Detection of the insertion in the XIP-I gene by PCR assays

8. Confirmation of the variation via microarray data analysis

9. Generation of Alternative Splicing Shift of Transcription Initiation Site Expansion of Intron Size Rearrangement of Promoter Regions Types of TE-Induced Genetic Variation

10. Identifying Transposon Insertion Polymorphisms between subspecies Comparative genomic method Homology-based method Structure-based method Step 1: Align Each Indica Contig (or BAC) to Japonica Genome Step 2: Find Cadidate Insertions and deletions between the two Genomes Step 3: Detect Transposon Insertions from Cadidate Insertions Step 4: Remove all artifacts via TSD Detection and Manual Checking

11. The Abundance of Transposon Insertion Polymorphisms in cultivated rice  2041 between Nipponbare and 93-11  691 between Nipponbare and Guangluai 4

12. SNP, TIP and Introgression Feltus et al, Genome Res. 14:1812-1819 (2004)

13. The Role on Intra-Variation in rice Outcome TIPs identified between Nip/93-11 TIPs identified between Nip/GLA4 TIPs estimated between rice subspecies Number Proportion (%) Number Proportion (%) Number a Proportion b (%) Alteration of cDNA sequence 874.3273.9449-4953.9-4.3 Rearrangement of promoter region 773.8162.3265-4382.3-3.8 Change of intron size 1095.3284.1472-6104.1-5.3 Total in the expressed gene region 27313.47110.31186-154310.3-13.4 Other TIPs176886.662089.79974-1033186.6-89.7 Total TIPs2041100.0691100.0~11517 c 100.0

14. A Global View of Transposon Insertion Polymorphisms in a collinear region

15. Majority of divergence between the two subspecies is due to TE insertions In Number In Size

16. TE insertion in NipponbareTE insertion in Guangluai 4 Copy no.Coverage(bp)Copy no.Coverage(bp) Class I Ty1/copia4627871235226678 Ty3/gypsy110953718127889838 Unclassified LTR231057812385415 Other Class I1026293839133 Total class I1891,364,5041931,241,064 Class II hAT13235411330542 En-Spm/CACTA231792081074619 MULE401337312784644 Tourist/Harbinger2166274111170 Mariner/Stowaway204394235251 Other MITE133941247562 Other Class II11787952544 Total class II141359,321143216,332 Other TEs1276751318568 Total TEs3421,731,5003491,475,964 Generating ~14% (53.5 Mb) of the genomic DNA sequence different between indica and japonica The smaller size of indica genome: Deletions of TEs occur more frequently

17. Retrotransposon Polymorphisms in Evolutionary Analysis PCR-based Assays Stable Polymorphisms In a class by oneself Ancestral State is known SSR DNA transposon polymorphism SNP ? ? ?

18. The Expected Pattern of Absence/Presence of Retrotransposon Insertion —— Insertion Event in Different Conditions

19. In Silico Analysis in ~23Mb Nip/GLA4 Collinear Region Distinct Evolutionary Route With Limited Introgression Variety Present in Nip Only Absent in GLA4 Only Not DeterminedTotal No. Nip+ 148 a (78.3%) + 15 (7.9%) + 26 (13.8%)189 (100%) GLA4--- 93-11-+ND Variety Absent in Nip Only Present in GLA4 Only Not DeterminedTotal No. Nip.- 100 b (51.8%) - 65 c (33.7%) - 28 (14.5%)193 (100%) GLA4+++ 93-11+-ND

20. Conclusions  Using an integrated approach, 2041 TIPs between Nip/93-11 and 691 TIPs between Nip/GLA4 were identified;  We estimated that there’re ~11517 Transposon Insertion Polymorphisms between indica and japonica;  They also generated ~14% of the genomic DNA (53.5 Mb) sequence different between subspecies;  > 10% of TE insertions affect more than 1000 genes via various ways;  Among them, the retrotransposon polymorphisms can be used in a wide range, like genetic diversity, evolutionary route and introgression of rice varieties.