Genetic Linkage Maps of Japanese and European Pears Aligned to the Apple Consensus Map

Transcription

1 Genetic Linkage Maps of Japanese and European Pears Aligned to the Apple Consensus Map T. Yamamoto 1, T. Kimura, T. aito 1, K. Kotobuki 1, N. Matsuta 1, R. Liebhard 3, C. Gessler 3, W.E. van de Weg and T. Hayashi 1 1 National Institute of Fruit Tree cience, Tsukuba, Ibaraki , Japan National Center for eeds and eedlings, Tsukuba, Ibaraki , Japan 3 wiss Federal Institute of Technology, 0 Zurich, witzerland Plant Research International, Wageningen 700 AA, The Netherlands Keywords: Pyrus pyrifolia, Pyrus communis, simple sequence repeats, linkage groups, synteny, Malus Abstract Genetic linkage maps of the Japanese pear (Pyrus pyrifolia Nakai) cultivar Housui and the European pear (Pyrus communis L.) cultivar Bartlett were constructed based on Amplified Fragment Length Polymorphism markers (AFLPs), imple equence Repeat markers (Rs) (from pear, apple and Prunus), isozymes, and phenotypic traits by using their F 1 progenies. The map of the female parent Bartlett consisted of 5 loci including AFLPs, 7 Rs (3 pear, 3 apple, 5 Prunus Rs), 1 isozyme and a self-incompatibility locus on 1 linkage groups over a total length of 0 cm. The map of Housui contained 0 loci including 1 AFLPs, Rs ( pear, apple, Prunus Rs), phenotypic traits and other markers on 0 linkage groups encompassing a genetic distance of 5 cm. These pear maps were aligned using 37 co-dominant markers that showed segregating alleles in both parents. Out of 0 tested R markers developed from apple, more than four-fifth could produce discrete amplified fragments in pear. Thirty-eight apple R markers showed 3 segregating loci in the linkage map of Bartlett, while 7 markers produced loci in Housui. All pear linkage groups could be successfully aligned to the apple consensus map by at least 1 apple Rs, suggesting that positions and linkages of R loci were well conserved between pear and apple. The selfincompatibility locus ( locus) was mapped to linkage group in Japanese and European pears as well as apple. Our results are the first major effort in comparative mapping of pear and apple. INTRODUCTION Pear (Pyrus spp.) is one of the most important fruits, which has been cultivated in Asia and Europe for 000 to 3000 years. Presently, pear is cultivated commercially in all the temperate regions in more than 50 countries around the world. However, there is very little information on genome research, i.e., genetic linkage maps, development of molecular markers, and genetic analysis on phenotypic traits. In this study, we constructed genetic linkage maps of the European pear cultivar Bartlett and the Japanese pear cultivar Housui using AFLP markers and R markers developed in pear, apple and Prunus. Both maps are aligned to the apple consensus map by using apple Rs as anchor loci. Genetic synteny between pear and apple is discussed. MATERIAL AND METHOD Plant Material An F 1 population of Bartlett (Pyrus communis L.) x Housui (P. pyrifolia Nakai) (3 individuals) was used for constructing genetic linkage maps. Genomic DNA was isolated from young leaves by a CTAB-based extraction method (Yamamoto et al., 00c). XI th Eucarpia ymp. on Fruit Breed. & Genetics Eds. F. Laurens and K. Evans Acta Hort. 3, IH 00 51

2 DNA Markers AFLP analysis was done with 0 sets obtained by Mse I primers (M/CAA, CAC, CAG, CAT, CTA, CTC, CTG, CTT) and 5 Eco RI primers labelled with FAM (E/AC, AG, TG, TC, GC). The AFLP products were separated and detected using a PRIM 377 DNA sequencer (PE Applied Biosystems). The size of the amplified bands was determined based on an internal standard DNA using Genecan software (PE Applied Biosystems). Fifty-four Rs developed from pear were used for the detection of microsatellite loci (Yamamoto et al., 00a, 00b, 00c), and were denoted by KA, HG, NB, NH, BG or RLG (Fig. 1, Fig. ). Eighty-four Rs developed in apple were screened and used for genetic mapping (Gianfranceschi et al., 1, Guilford et al., 17, Liebhard et al., 00). A total of 5 R markers developed in Prunus (peach, cherry) were also tested. R amplification was performed as described by Yamamoto et al. (00c). Linkage Analysis JoinMap ver..0 was used for the construction of genetic linkage maps of Bartlett and Housui. The Kosambi function was used to convert recombination units into genetic distances. The mapping analysis was conducted using a minimum LOD score of.0. The same numbering system of linkage groups was performed as that of apple (Maliepaard et al., 1) by using common apple Rs as anchor loci (Liebhard et al., 00). REULT AND DICUION Genetic Linkage Maps of Japanese and European Pears The genetic linkage map of Bartlett was constructed with 5 DNA markers, including 3 apple Rs, 3 pear Rs, 5 Prunus Rs, 1 isozyme, 1 locus and AFLPs (Fig. 1). The obtained map consisted of 1 linkage groups that covered more than 0 cm with an average distance of.0 cm between each pair of loci. The size of the linkage groups ranged from cm (Linkage Group (LG)) to cm (LG). The segregation of almost all R markers fitted the expected ratio of monogenic inheritance. The segregation of many markers on LG1 was largely distorted. The self-incompatibility locus ( locus) was found in bottom of LG. The genetic linkage map of Housui contained 0 loci (1 AFLPs, pear Rs, apple Rs, Prunus Rs, phenotypic traits and other markers) shown in Fig.. The map consisted of 0 linkage groups encompassing a genetic distance of 5 cm. Genetic linkage maps of these pears were aligned using 37 co-dominant markers that showed segregating alleles in both parents. ynteny between Pear and Apple Thirty-six Rs originating from apple were mapped in genetic linkage maps of the European pear Bartlett and the apple consensus map (Table 1). Only R loci (CH01b1, CH0d0) were assigned to different linkage groups between pear and apple. The other 3 apple R loci could be positioned in presumably homologous linkage groups of pear. All pear linkage groups could be successfully aligned to the apple consensus map by at least 1 apple R (or isozyme AAT-1 for LG), suggesting that positions and linkages of R loci were well conserved between them, and that the genome organization of apple and pear is highly similar. The self-incompatibility locus was found at the bottom of LG in Japanese and European pears as well as apple. Our results are the first major effort in comparative mapping of pear and apple. Literature Cited Gianfranceschi, L., eglias, N., Tarchini, R., Komjanc, M. and Gessler, C. 1. imple sequence repeats for the genetic analysis of apple. Theor. Appl. Genet. :-7. Guilford, P., Prakash,., Zhu, J.M., Rikkerink, E., Gardiner,., Bassett, H. and Forster, R. 17. Microsatellites in Malus x domestica (apple): abundance, polymorphism and 5

3 cultivar identification. Theor. Appl. Genet. :-5. Maliepaard, C., Alston, F.H., van Arkel, G., Brown, L.M., Chevreau, E., Dunemann, F., Evans, K.M., Gardiner,., Guilford, P., van Heusden, A.W., Janse, J., Laurens, F., Lynn, J.R., Manganaris, A.G., den Nijs, A.P.M., Periam, N., Rikkerink, E., Roche, P., Ryder, C., ansavini,., chmidt, H., Tartarini,., Verhaegh, J.J., Vrielink-van Ginkel, M. and King, G.J. 1. Aligning male and female linkage maps of apple (Malus pumila Mill.) using multi-allelic markers. Theor. Appl. Genet. 7:0-73. Liebhard, R., Gianfranceschi, L., Koller, B., Ryder, C.D., Tarchini, R., van de Weg, E. and Gessler, C. 00. Development and characterisation of new microsatellites in apple (Malus x domestica Borkh.). Mol. Breed. :-1. Yamamoto, T., Kimura, T., awamura, Y., Manabe, T., Kotobuki, K., Hayashi, T., Ban, Y. and Matsuta, N. 00a. imple sequence repeats for genetic analysis in pear. Euphytica 1: 1-7. Yamamoto, T., Kimura, T., hoda, M., Ban, Y., Hayashi, T. and Matsuta, N. 00b. Development of microsatellite markers in Japanese pear (Pyrus pyrifolia Nakai). Molecular Ecology Notes :1-. Yamamoto, T., Kimura, T., hoda, M., Imai, T., aito, T., awamura, Y., Kotobuki, K., Hayashi, T. and Matsuta, N. 00c. Genetic linkage maps constructed by using an interspecific cross between Japanese and European pears. Theor. Appl. Genet. :-. 53

4 Tables Table 1. Mapped linkage group of apple R loci in genetic maps of pear and apple. Linkage group R Locus Pear (Bartlett) Apple a CH05g0 #1- #1 CH03g1 #1,3 #1,3 CH0b # # CH0f0 # # M1h03 #3 #3 CH0d0 #3 # NZ05g # # CH0c0b # # M0c0 # CH0b1 #5 #5, CH05e0 #5 #5 CH03d1 # # # # CH0e05 #7 #7 # # CH01h0 # #, CH0b03b # # # # CH0c # # CH0h0 # # CH03d0 # # CH0g07 # # NZf #1 #1 CH01f0 #1 #1 CH0d0 #1 #1 CH05d0 #1 #1 CH03a0 # # CH05f0 # # CH01g05 #1 #1 M01a05 #1 #1 CH03g0 #1 #1 NZ0b1 #15 #15 CH0d #15 #15 #15 #15 CH0c0 #15 #15 CH05c0 # # CH01b1 # #,1, # # a Liebhard et al.,003; van de Weg (pers. commun.) 5

5 55 Figuress Fig. 1. A genetic linkage map of the European pear Bartlett. Genetic distances and loci are listed on the left and the right sides, respectively. The designation of AFLP markers is based on primer combination and size. R loci from apple and pear are underlined and in italics, respectively. The self-incompatibility locus is denoted by on LG. Asterisks indicate distorted segregations of markers according to the chi-square test. Distortions at the 5%, 1% and 0.1% level are indicated as *, ** and ***, respectively. RLG AG/CAA-7 5. TG/CTC-3. AC/CAC-7 AG/CAT P1A0- CH03g1- TC/CTG-35. GC/CTC-5 1 GC/CTC RLG AG/CAA-7 5. TG/CTC-3. AC/CAC-7 AG/CAT P1A0- CH03g1- TC/CTG-35. GC/CTC-5 1 GC/CTC AC/CAT-37 AC/CTG- KAb.3 TC/CAG- 1. NH0a 1.3 CH05g0. 1- AC/CAT-37 AC/CTG- KAb.3 TC/CAG- 1. NH0a 1.3 CH05g0. 1- CH0f0 AC/CAC-5.5 GC/CAC-0 GC/CAC-. AC/CTT-5. BGT3b 3.0 GC/CTT GC/CTA-77* 3. TG/CTC CH0b 7.1 AG/CTC- AC/CAG-5.7 NH00b 0. TC/CTA-30.3 CH0f0 AC/CAC-5.5 GC/CAC-0 GC/CAC-. AC/CTT-5. BGT3b 3.0 GC/CTT GC/CTA-77* 3. TG/CTC CH0b 7.1 AG/CTC- AC/CAG-5.7 NH00b 0. TC/CTA-30.3 AC/CTA-3 AC/CAG-0. UDP-.0 CH0d0. NH03a.3 TG/CAA-3.3 TC/CAT AG/CTG- 37. AC/CTG-31.5 M1h03 GC/CAA-5 CH03g TG/CAC AC/CTA-3 AC/CAG-0. UDP-.0 CH0d0. NH03a.3 TG/CAA-3.3 TC/CAT AG/CTG- 37. AC/CTG-31.5 M1h03 GC/CAA-5 CH03g TG/CAC AG/CAT-70 TG/CAA M0c0 TC/CAA-30*. AC/CAG-150. GC/CAC-373. NZ05g AG/CTC- NH0a 3.7 AC/CTG-7 TG/CAA TC/CAC-1 AC/CAG TC/CAG-3 TC/CAG TG/CAC- TC/CAT CH0c0b AC/CAG-3 TG/CTT AG/CTG- 7.5 AG/CAT-70 TG/CAA M0c0 TC/CAA-30*. AC/CAG-150. GC/CAC-373. NZ05g AG/CTC- NH0a 3.7 AC/CTG-7 TG/CAA TC/CAC-1 AC/CAG TC/CAG-3 TC/CAG TG/CAC- TC/CAT CH0c0b AC/CAG-3 TG/CTT AG/CTG- 7.5 AG/CAT-71 TG/CAT- 0. AG/CAC- TC/CTG-303 AG/CTA-77 TC/CAA-5. AG/CAT-30 AG/CAA CH0b1.3 AC/CAA-30.0 NH00a-1 NB3a AG/CAT-331. AC/CTA-.0 GC/CAA-* 5. AC/CAC-3.7 TG/CTG CH05e0* 55. NB1a 5. AC/CAG-1. GC/CTG-353. TG/CAG AG/CAT-71 TG/CAT- 0. AG/CAC- TC/CTG-303 AG/CTA-77 TC/CAA-5. AG/CAT-30 AG/CAA CH0b1.3 AC/CAA-30.0 NH00a-1 NB3a AG/CAT-331. AC/CTA-.0 GC/CAA-* 5. AC/CAC-3.7 TG/CTG CH05e0* 55. NB1a 5. AC/CAG-1. GC/CTG-353. TG/CAG AC/CTG-5 AG/CAG- AC/CAG- 1. GC/CAC AC/CAG CH03d1 3. TC/CAC-1 TC/CTA-77.5 TG/CTG-3 5. AC/CTG-5 AG/CAG- AC/CAG- 1. GC/CAC AC/CAG CH03d1 3. TC/CAC-1 TC/CTA-77.5 TG/CTG-3 5. AC/CAT- TC/CTA-7 TG/CTT-3.1 TG/CAC-1 1 GC/CAG-1. CH0e05 0. TG/CTG-. 7 AC/CAT- TC/CTA-7 TG/CTT-3.1 TG/CAC-1 1 GC/CAG-1. CH0e05 0. TG/CTG-. 7 AAT-1 TG/CTG-7.0 TG/CTG-. AG/CAC TC/CTC-.5 AAT-1 TG/CTG-7.0 TG/CTG-. AG/CAC TC/CTC-.5 NB1a-1 AG/CTA- TC/CAG-5 NH0a.7 AG/CAA-75.1 AC/CAG-.1 KA0.7 GC/CTC-. NBa 35.3 TG/CTG-* 1. CH01h0* TC/CAG TC/CTC-0* NB1a-1 AG/CTA- TC/CAG-5 NH0a.7 AG/CAA-75.1 AC/CAG-.1 KA0.7 GC/CTC-. NBa 35.3 TG/CTG-* 1. CH01h0* TC/CAG TC/CTC-0* GC/CAT-15 AC/CAG-3. M7a 15.3 AC/CAT-7 1. AC/CAG AC/CAA-. GC/CTG AC/CAC-3 1. TC/CAA-37.1 CH0c. GC/CAT GC/CAG-. CH0b03b GC/CAT-15 AC/CAG-3. M7a 15.3 AC/CAT-7 1. AC/CAG AC/CAA-. GC/CTG AC/CAC-3 1. TC/CAA-37.1 CH0c. GC/CAT GC/CAG-. CH0b03b GC/CTA-* CH0h0. NB1a-. AC/CAC-5. CH03d0 AC/CTG-15. AG/CAC AC/CAC-1 3. AC/CAC- AC/CAG-0.5 GC/CTC AC/CTA- 5. AC/CTA-15 NB5a CH0g07 GC/CTA-3 7. AC/CTA GC/CTA-* CH0h0. NB1a-. AC/CAC-5. CH03d0 AC/CTG-15. AG/CAC AC/CAC-1 3. AC/CAC- AC/CAG-0.5 GC/CTC AC/CTA- 5. AC/CTA-15 NB5a CH0g07 GC/CTA-3 7. AC/CTA TC/CTG-55* TG/CAC-3 CH05d0. NZf.1 TC/CTA-7 AC/CAG-07 KA TG/CTT AG/CAG AG/CAC-7. AG/CTA GC/CAG- CH01f0 0. CH0d TC/CTG-55* TG/CAC-3 CH05d0. NZf.1 TC/CTA-7 AC/CAG-07 KA TG/CTT AG/CAG AG/CAC-7. AG/CTA GC/CAG- CH01f0 0. CH0d TG/CTG-1 GC/CTG-7 TC/CTA-0 NH00b GC/CTT- 35. TC/CTG AC/CTC-5 3. CH03a0 1. TC/CTA- TC/CAC-0 GC/CTA-0 GC/CTT-1 3. AC/CTG-30 CH05f0 NH01a AG/CTA-.5 TG/CTG-3 5 TG/CTG-1 GC/CTG-7 TC/CTA-0 NH00b GC/CTT- 35. TC/CTG AC/CTC-5 3. CH03a0 1. TC/CTA- TC/CAC-0 GC/CTA-0 GC/CTT-1 3. AC/CTG-30 CH05f0 NH01a AG/CTA-.5 TG/CTG-3 5 NH00a TC/CTG-3*** 1.0 GC/CTA-*** AG/CTC-*** 0.7 GC/CTT-*** 7. CH01g05*** M01a05*** 35. GC/CTT-0*** 3 NH001c*** 3. GC/CTT-*** 1.1 CH03g0***. AC/CTT- 7 1 NH00a TC/CTG-3*** 1.0 GC/CTA-*** AG/CTC-*** 0.7 GC/CTT-*** 7. CH01g05*** M01a05*** 35. GC/CTT-0*** 3 NH001c*** 3. GC/CTT-*** 1.1 CH03g0***. AC/CTT- 7 1 TG/CAC- NH07a 0. NZ0b1 GC/CTG- 3. P1A0-1.7 NH05a. TC/CAC-.0 TC/CTG-15 AC/CAC-303. TC/CTC GC/CAC-. AG/CTA- CH0d* 5. GC/CAT- 5. CH0c0 7 TG/CTC TG/CAC- NH07a 0. NZ0b1 GC/CTG- 3. P1A0-1.7 NH05a. TC/CAC-.0 TC/CTG-15 AC/CAC-303. TC/CTC GC/CAC-. AG/CTA- CH0d* 5. GC/CAT- 5. CH0c0 7 TG/CTC KA1 CH05c0-1.0 NH007b 5. Ma AC/CTA-30 AC/CAG AG/CAG- AG/CAC-1 1 TC/CAT-3. TG/CTG- 35. AC/CTC-5.3 AC/CTG-0.3 GC/CAT-0 GC/CAC-7 AC/CAC KA1 CH05c0-1.0 NH007b 5. Ma AC/CTA-30 AC/CAG AG/CAG- AG/CAC-1 1 TC/CAT-3. TG/CTG- 35. AC/CTC-5.3 AC/CTG-0.3 GC/CAT-0 GC/CAC-7 AC/CAC TG/CAG-15 NB1a- 5. GC/CAA-3. GC/CAA-35 AC/CAG- TC/CAG AG/CAC- 0. AC/CTA- 1. CH01b AC/CTA AG/CTG NH00b-1 GC/CTG-5.7 TC/CTG TG/CAG- 50. TG/CTC TC/CTC- 5. TG/CAG-15 NB1a- 5. GC/CAA-3. GC/CAA-35 AC/CAG- TC/CAG AG/CAC- 0. AC/CTA- 1. CH01b AC/CTA AG/CTG NH00b-1 GC/CTG-5.7 TC/CTG TG/CAG- 50. TG/CTC TC/CTC- 5. AG/CTG-7 TC/CAA-.0 TC/CAT-1.1 TC/CAG-03 AG/CTG-7 TC/CAA-.0 TC/CAT-1.1 TC/CAG-03 AG/CTG-7 TC/CAA-.0 TC/CAT-1.1 TC/CAG-03

6 5 Fig.. A genetic linkage map of the Japanese pear Housui. Genetic distances and loci are listed on the left and the right sides, respectively. The self-incompatibility locus and the young leaf color are denoted by and Lc on LG, respectively. CH0h0 GC/CAG-7* GC/CTT-1*. CH03d0.5 NB1a AC/CTC-07.0 AC/CAA-3. GC/CAG AC/CAC TC/CAT RLG1-3. GC/CAC- 5. GC/CTC- NB0b-1.7 CH0g AC/CTA-7.7 TG/CAA-* 1. TC/CTT-* 1. BGT3*** AC/CTG-* TC/CTT-3* 1.1 GC/CTG-31* 0.7 AC/CAG-*.0 CH0b 3. TC/CTA AG/CAG-15 TC/CAG-5 TG/CTT-55* TG/CTC AC/CAC-3.0 AC/CAT- 0. TC/CTT-7 NH0a 3.7 NH005b 3.1 AC/CAA- TC/CTG-3 AC/CTA-5.3 CH01h.1 NH00a- NH0a. AC/CTA-150 AG/CAG-1. CH0c 3. CH03d BGT 0.5 GC/CTT CH0b03b. GC/CAG-7 AC/CTT- 7. GC/CAG-0.7 AC/CAA-3 P1A0-3.3 CH03g1-3 TC/CAT TC/CAC- 1. TG/CAT-3 1. AG/CTC-5 IM 5. KAb TC/CTC TG/CAC-7 TC/CAC-7 5. TC/CTT TC/CAC-370 TC/CAC-33 CH0d0. NH03a.7 TG/CAG-.5 AC/CTT-370* 3-1 NH030a 1. TG/CAA-. TC/CAG-31.1 GC/CTA-71 NB0b-. M1h03 1. HGAb. CH03g CH0c0b TG/CAC- TG/CAA- GC/CAG AG/CAG UDP TG/CTT-35 AG/CAT-3. GC/CAT-3 3. TC/CAA CH03d1 5. TG/CAC- 5.7 AG/CAG AG/CAC AC/CAG-1 UDP-01 NH00b-3 5. GC/CAA CH01h0 1.5 TG/CAC-7.1 AG/CAC- 5.1 AC/CTG-7 TC/CAT-3 GC/CAA-1 1. GC/CAC-53 CH05c0- AG/CAT-5* 3.7 AC/CAG- Ma NH00a* AG/CTG-. CH01g05 AG/CAA GC/CTA-00 3 GC/CAC-0 TC/CAG-0 5. M01a05 AC/CTA-0 1. CH03g AG/CAG-3 GC/CAC- P1A TC/CAT- NH05a 3. TC/CAC TC/CTT CH0d AC/CAA-7 CH01e GC/CAC- 5.1 UDP-01-1 TG/CTC-31 AC/CAC-1 3. CH0c0 NBa.1 TG/CTC AG/CAG-1 CH05c NH0a GC/CTA-3. TG/CAA TC/CAT- CH05g GC/CTC HGT. GC/CAT-335 AC/CTA- 1. AC/CTC-.3 AC/CTG-0 3 Lc 1. AG/CTG- 5.5 AG/CAC- NH00b- NH00b AG/CAG-.0 GC/CAC-1 AG/CAG-. AC/CTT-35* TC/CTG-3* 7.3 MDH- DIA-1 DIA- 1 GC/CTG RLG1- AG/CAC-0 NH01b AG/CAC AC/CAG-0 GC/CAA-. 0 CH0h0 GC/CAG-7* GC/CTT-1*. CH03d0.5 NB1a AC/CTC-07.0 AC/CAA-3. GC/CAG AC/CAC TC/CAT RLG1-3. GC/CAC- 5. GC/CTC- NB0b-1.7 CH0g AC/CTA-7.7 TG/CAA-* 1. TC/CTT-* 1. CH0h0 GC/CAG-7* GC/CTT-1*. CH03d0.5 NB1a AC/CTC-07.0 AC/CAA-3. GC/CAG AC/CAC TC/CAT RLG1-3. GC/CAC- 5. GC/CTC- NB0b-1.7 CH0g AC/CTA-7.7 TG/CAA-* 1. TC/CTT-* 1. BGT3*** AC/CTG-* TC/CTT-3* 1.1 GC/CTG-31* 0.7 AC/CAG-*.0 CH0b 3. TC/CTA AG/CAG-15 BGT3*** AC/CTG-* TC/CTT-3* 1.1 GC/CTG-31* 0.7 AC/CAG-*.0 CH0b 3. TC/CTA AG/CAG-15 TC/CAG-5 TG/CTT-55* TG/CTC AC/CAC-3.0 AC/CAT- 0. TC/CTT-7 NH0a 3.7 NH005b 3.1 AC/CAA- TC/CTG-3 AC/CTA-5.3 CH01h.1 TC/CAG-5 TG/CTT-55* TG/CTC AC/CAC-3.0 AC/CAT- 0. TC/CTT-7 NH0a 3.7 NH005b 3.1 AC/CAA- TC/CTG-3 AC/CTA-5.3 CH01h.1 NH00a- NH0a. AC/CTA-150 AG/CAG-1. CH0c 3. CH03d BGT 0.5 GC/CTT CH0b03b. GC/CAG-7 AC/CTT- 7. GC/CAG-0.7 NH00a- NH0a. AC/CTA-150 AG/CAG-1. CH0c 3. CH03d BGT 0.5 GC/CTT CH0b03b. GC/CAG-7 AC/CTT- 7. GC/CAG-0.7 AC/CAA-3 P1A0-3.3 CH03g1-3 TC/CAT TC/CAC- 1. TG/CAT-3 1. AG/CTC-5 IM 5. AC/CAA-3 P1A0-3.3 CH03g1-3 TC/CAT TC/CAC- 1. TG/CAT-3 1. AG/CTC-5 IM 5. KAb TC/CTC KAb TC/CTC TG/CAC-7 TC/CAC-7 5. TC/CTT TC/CAC-370 TC/CAC-33 CH0d0. NH03a.7 TG/CAG-.5 AC/CTT-370* 3-1 TG/CAC-7 TC/CAC-7 5. TC/CTT TC/CAC-370 TC/CAC-33 CH0d0. NH03a.7 TG/CAG-.5 AC/CTT-370* 3-1 NH030a 1. TG/CAA-. TC/CAG-31.1 GC/CTA-71 NB0b-. M1h03 1. HGAb. CH03g NH030a 1. TG/CAA-. TC/CAG-31.1 GC/CTA-71 NB0b-. M1h03 1. HGAb. CH03g CH0c0b TG/CAC- CH0c0b TG/CAC- TG/CAA- GC/CAG AG/CAG UDP TG/CTT-35 AG/CAT-3. GC/CAT-3 3. TC/CAA CH03d1 5. TG/CAC- 5.7 AG/CAG AG/CAC TG/CAA- GC/CAG AG/CAG UDP TG/CTT-35 AG/CAT-3. GC/CAT-3 3. TC/CAA CH03d1 5. TG/CAC- 5.7 AG/CAG AG/CAC AC/CAG-1 UDP-01 NH00b-3 5. GC/CAA CH01h0 1.5 TG/CAC-7.1 AG/CAC- 5.1 AC/CTG-7 AC/CAG-1 UDP-01 NH00b-3 5. GC/CAA CH01h0 1.5 TG/CAC-7.1 AG/CAC- 5.1 AC/CTG-7 TC/CAT-3 GC/CAA-1 1. GC/CAC-53 CH05c0- AG/CAT-5* 3.7 AC/CAG- TC/CAT-3 GC/CAA-1 1. GC/CAC-53 CH05c0- AG/CAT-5* 3.7 AC/CAG- Ma NH00a* AG/CTG-. CH01g05 AG/CAA GC/CTA-00 3 GC/CAC-0 TC/CAG-0 5. M01a05 AC/CTA-0 1. CH03g Ma NH00a* AG/CTG-. CH01g05 AG/CAA GC/CTA-00 3 GC/CAC-0 TC/CAG-0 5. M01a05 AC/CTA-0 1. CH03g AG/CAG-3 GC/CAC- P1A TC/CAT- NH05a 3. TC/CAC TC/CTT AG/CAG-3 GC/CAC- P1A TC/CAT- NH05a 3. TC/CAC TC/CTT CH0d AC/CAA-7 CH01e GC/CAC- 5.1 UDP-01-1 TG/CTC-31 AC/CAC-1 3. CH0c0 NBa.1 TG/CTC CH0d AC/CAA-7 CH01e GC/CAC- 5.1 UDP-01-1 TG/CTC-31 AC/CAC-1 3. CH0c0 NBa.1 TG/CTC AG/CAG-1 CH05c NH0a GC/CTA-3. AG/CAG-1 CH05c NH0a GC/CTA-3. TG/CAA TC/CAT- CH05g GC/CTC HGT. GC/CAT-335 AC/CTA- 1. AC/CTC-.3 AC/CTG-0 3 Lc 1. AG/CTG- 5.5 AG/CAC- NH00b- NH00b AG/CAG-.0 GC/CAC-1 AG/CAG-. AC/CTT-35* TC/CTG-3* 7.3 TG/CAA TC/CAT- CH05g GC/CTC HGT. GC/CAT-335 AC/CTA- 1. AC/CTC-.3 AC/CTG-0 3 Lc 1. AG/CTG- 5.5 AG/CAC- NH00b- NH00b AG/CAG-.0 GC/CAC-1 AG/CAG-. AC/CTT-35* TC/CTG-3* 7.3 MDH- DIA-1 DIA- 1 GC/CTG MDH- DIA-1 DIA- 1 GC/CTG RLG1- AG/CAC-0 NH01b AG/CAC RLG1- AG/CAC-0 NH01b AG/CAC AC/CAG-0 GC/CAA-. 0 AC/CAG-0 GC/CAA-. 0