IDENTIFICATION OF QTLS FOR STARCH CONTENT IN SWEETPOTATO (IPOMOEA BATATAS (L.) LAM.)

Transcription

1 Journal of Integrative Agriculture Advanced Online Publication: 2013 Doi: /S (13) IDENTIFICATION OF QTLS FOR STARCH CONTENT IN SWEETPOTATO (IPOMOEA BATATAS (L.) LAM.) YU Xiao-xia 1*, ZHAO Ning 1*, LI Hui 1*, JIE Qin 1, ZHAI Hong 1, HE Shao-zhen 1, LI Qiang 2 and LIU Qing-chang 1** 1 Laboratory of Crop Heterosis and Utilization, Ministry of Education/Beijing Key Laboratory of Crop Genetic Improvement/China Agricultural University, Beijing , P.R.China 2 Xuzhou Sweetpotato Research Center, Xuzhou , P.R.China * These authors contributed equally to this work. ** Correspondence LIU Qing-chang, Tel/Fax: , liuqc@cau.edu.cn Abstract Sweetpotato (Ipomoea batatas (L.) Lam.) breeding is challenging due to its genetic complexity. In the present study, interval mapping (IM) and multiple quantitative trait locus (QTL) model (MQM) analysis were used to identify QTLs for starch content with a mapping population consisting of 202 F 1 individuals of a cross between Xushu 18, a cultivar susceptible to stem nematodes, with high yield and moderate starch, and Xu 781, which is resistant to stem nematodes, has low yield and high starch content. Six QTLs for starch content were mapped on six linkage groups of the Xu 781 map, explaining % of the variation. Especially, one of them, DMFN_4, accounted for 38.8% of starch content variation, which is the QTL that explains highest phenotypic variation detected to date in sweetpotato. All of the six QTLs had a positive effect on the variation of the starch content, which indicated the inheritance derived from the parent Xu 781. Two QTLs for starch content were detected on two linkage groups of the Xushu 18 map, explaining 14.3 and 16.1% of the variation, respectively. They had a negative effect on the variation, indicating the inheritance derived from Xu 781. Seven of eight QTLs were co-localized with a single marker. This is the first report on the development of QTLs co-localized with a single marker in sweetpotato. These QTLs and their co-localized markers may be used in marker-assisted breeding for the starch content of sweetpotato. Key words: co-localized marker, Ipomoea batatas (L.) Lam., QTLs, starch content INTRODUCTION Sweetpotato, Ipomoea batatas (L.) Lam., is an important food and industrial material crop in the world. It is also an alternative source of bio-energy as a raw material for fuel production (Zang et al. 2009). This crop has high starch content, which is used as an ideal material of starch products and fuel ethanol (Yamakawa 1998). Therefore, much breeding work in sweetpotato is focused on the development of cultivars with high starch content (CIP 2010). Sweetpotato breeding is challenging due to its genetic complexity. Most traits of economical importance, including starch content, yield and many resistance traits, are quantitatively inherited or appear to be so due to the

2 polyploid nature of this crop (Cervantes-Flores et al. 2008). Marker-assisted breeding tool is needed for sweetpotato. Therefore, the mapping of quantitative trait loci (QTLs) associated to these important quantitative traits is necessary. To date, there are only a few reports on the mapping of QTLs in sweetpotato. Wu et al. (2005) detected one QTL for starch content using a mapping population consisting of 46 F 1 individuals of a cross between Mianfen 1 with high starch content and Hongqi 4 with low starch content. However, the QTL was not accurate and stable due to the small mapping population (46 individuals) and low-density genetic map with only 21 sequence related amplified polymorphism (SRAP) markers. Li et al. (2010) identified one QTL for starch content, explaining only 7.7% of the variation, using a mapping population consisting of 240 F 1 individuals of a cross between Luoxushu 8 with high starch content and Zhengshu 20 with low starch content. Using the mapping population consisting of 240 progeny derived from a cross between an African cultivar Tanzania and an American cultivar Beauregard, Cervantes-Flores et al. (2011) located 13 QTL for storage root dry-matter content, 12 QTL for starch content and 8 QTL for ß-carotene content, explaining 15-24% of the variation in dry-matter content, 17-30% of the starch content and 17-35% of ß-carotene content. We developed a high-density linkage map of sweetpotato based on amplified fragment length polymorphism (AFLP) and expressed sequence tag-simple sequence repeat (EST-SSR) markers using a mapping population consisting of 202 F 1 individuals of a cross between Xushu 18 and Xu 781, which are the core parents widely used in sweetpotato breeding of China (Zhao 2012). In the present study, using this mapping population, we mapped 8 QTLs for starch content, explaining 9.1% to 38.8% of the variation in starch content. RESULTS Starch content analysis The starch content of the mapping population was distributed normally based on each year and mean data (Fig. 1). The average starch content of the mapping population in three years ranged from 8.5 to 23.8% with a population mean of 17.6%. The average starch content of Xushu 18 and Xu 781 was 14.2 and 21.8%, respectively. Analysis of variance (ANOVA) showed that no significant difference of starch content existed among 3 yr. Therefore, data from the three years were averaged for the QTLs analysis. Transgressive segregation was also observed in the population, with some progenies exhibiting higher levels and others lower levels of starch content than either parent. QTL mapping The QTLs for starch content detected based on each year and the mean data were listed in Table 1. Based on the mean of starch content in three years, six QTLs for starch content, named DMFN_1, DMFN_2, DMFN_3, DMFN_4, DMFN_5, and DMFN_6, respectively, were mapped on six linkage groups of Xu 781, XU781(01.05), XU781(02.07), XU781(03.15), XU781(07.40), XU781(08.45) and XU781(10.54), and explained % of the variation in starch content (Fig. 2). One major QTL DMFN_4 located on XU781(07.40) accounted for 38.8% of starch content variation with a 2

3 peak LOD score of 7.2. This is the QTL which explains highest phenotypic variation detected to date in sweetpotato. Another major QTL DMFN_1 on XU781(01.05) explained 20.5 % of the variation with a peak LOD score of 5.0. Four QTLs, DMFN_2 on XU781(02.07), DMFN_5 on XU781(08.45), DMFN_3 on XU781(03.15) and DMFN_6 on XU781(10.54), explained 14.4, 14.0, 10.6, and 9.1% of the variation, respectively. Five of the six QTLs, DMFN_1, DMFN_2, DMFN_3, DMFN_5, and DMFN_6, were co-localized with a single marker, E31M19-21s**, E51M28-6d, E17M55-18d, E50M49-3d and C61_2d, respectively (Table 1). The remaining one QTL, DMFN_4, was located between E27M62-2d and E36M63-10ds** (Table 1). Two QTLs for starch content, named DMMN_1 and DMMN_2, respectively, were detected on two linkage groups of the Xushu 18 map, explaining 14.3% and 16.1% of the variation in starch content, respectively. These two QTLs were co-localized with a single marker, E1M31-17ds** and E50M12-8s, respectively. All of the eight QTLs were detected in at least two environments and the mean data and were rather stable (Table 1). All of the six QTLs located on the Xu 781 map had a positive effect on the variation of the starch content, while the two QTLs on the Xushu 18 map had a negative effect on the variation, indicating their inheritance derived from Xu 781. These results agree with the fact that Xu 781 has the high starch content. Thus, these QTLs and their co-localized markers may be used for marker-assisted selection of starch content in sweetpotato. DISCUSSION The need for improved breeding tools to facilitate sweetpotato improvement is great due to its genetic complexity. Most traits of economical importance are quantitatively inherited (Cervantes-Flores et al. 2008). The mapping of QTLs associated to these important quantitative traits is needed for marker-assisted breeding of sweetpotato. To the best of our knowledge, the QTL mapping study is very poor in sweetpotato. Cervantes-Flores et al. (2008) developed linkage maps of Tanzania and Beauregard, and both maps consisted of 86 and 90 linkage groups with 1166 and 960 AFLP markers, respectively. Cervantes-Flores et al. (2011) identified some QTLs for storage root dry-matter content, starch content and ß-carotene content. China is the largest producer of sweetpotato with the planting area of 4.5 million ha -1 annually, accounting for approximately 50% of the world. Li et al. (2010) identified one QTL for starch content, explaining only 7.7% of the variation, using a mapping population of the cross Luoxushu 8 Zhengshu 20. In our previous study, a high-density linkage map was constructed based on AFLP and EST-SSR markers by the use of a mapping population consisting of 202 F 1 population of a cross between Xushu 18 and Xu 781 (Zhao 2012). Xushu 18 map included 1936 AFLP and 141 EST-SSR markers, and Xu 781 map contained 1824 AFLP and 130 EST-SSR markers (Zhao 2012). In the present study, eight QTLs for starch content were mapped on the maps of Xu 781 and Xushu 18, and explained 9.1 to 38.8% of the variation in starch content. Especially, one of them, DMFN_4 located on XU781(07.40) accounted for 38.8% of starch content variation, which is the QTL that explains highest phenotypic variation reported to date in sweetpotato. In addition, seven of the eight QTLs were co-localized with a single marker. This is the first report on the development of QTLs co-localized with a single marker in sweetpotato. Transgressive segregation was also observed for starch content in the progeny of the mapping population. One possible reason is that the high levels of heterozygosity of the parents caused the accumulation or loss of 3

4 certain favorable alleles in the progeny (Tanksley 1993; Young 1996; Cervantes-Flores et al. 2011). It is a usual phenomenon in applied breeding program of sweetpotato, especially when the crosses are made between distinct genetic materials. CONCLUSION Eight QTLs for starch content, explaining % of the variation, have been mapped on the linkage maps of Xu 781 and Xushu 18. The inheritance of all the QTLs is derived from Xu 781 with high starch content. Seven of eight QTLs were co-localized with a single marker. Thus, these QTLs and their co-localized markers may be used in marker-assisted breeding for the starch content of sweetpotato. MATERIALS AND METHODS Plant materials Sweetpotato cultivar Xushu 18 and line Xu 781 were employed in this study. Xushu 18 is a cultivar, which is susceptible to stem nematodes, with high yield and moderate starch, and is widely planted in China. Xu 781 is resistant to stem nematodes, has low yield and high starch content, and is the core parent widely used in sweetpotato breeding of China. An F 1 population consisting of 202 F 1 individuals of a cross between Xushu 18 and Xu 781 was used as the mapping population. Field design The 202 F 1 individuals and both parents were planted at Langfang Experimental Station of Langfang Academy of Agriculture and Forestry Sciences, Hebei, China, during 2009 to The field trial was arranged in randomized complete block design (RCBD) with three replications per experiment. Each plot contained 20 plants, which were grown in two rows spacing 25 cm within rows and 80 cm between rows. All of the experiments were conducted under the normal field conditions. Approximately 120 days after planting the experiments were harvested. Starch content determination Starch content was determined based on dry-matter content of storage roots. Using a dry weight conversion method, dry-matter content was measured by the percentage of dry weight to the fresh weight of harvested storage roots. There were five medium-sized storage roots randomly sampled per plot for the following measurement at harvest. The sampled storage roots were washed up and sliced into approximately 4 mm chips with a food processor. A mixed fresh sample of slices was weighed 100 g and oven-dried at 105 C for 30 min, then 60 C for 5 h, at last 80 C for 48 h. The same procedures were followed for all replications for a total of 1836 samples ((202 individuals+2 parents) 3 replications 3 years). The conversion formula of the starch content in sweetpotato described by Wang et al. (1989) is followed, y= x , in which y is starch content and x is dry-matter content. 4

5 QTL analysis The construction of Xushu 18 Xu 781 genetic maps was described by Zhao (2012). Xushu 18 map included 1936 AFLP and 141 EST-SSR markers, and Xu 781 map contained 1824 AFLP and 130 EST-SSR markers (Zhao 2012). The general linear model (GLM) procedure of SAS v9.1 (Cary, NC, USA) was used to perform ANOVA of starch content. The average starch content data of all three years in each environment were used to the subsequent QTL detection. Interval mapping (IM) was performed first and, subsequently, multiple QTL model (MQM) was used to refine the position and magnitude of QTL with MapQTL software 4.0 (Van Ooijen 2004 ). In MQM analysis, the marker closest to the QTLs was considered as a cofactor. The presence of different QTLs was determined by the threshold of a LOD score of 2.5. The QTLs detected at the same genomic location in the mean environment and at least two different environments were determined as a single QTL. The final QTL mapping was drawn using Map Chart 2.2 (Voorrips 2002). Acknowledgements This work was supported by China Agriculture Research System (CARS-11, Sweetpotato) and the National 863 project of China (2012AA101204). References Cervantes-Flores J C, Sosinski B, Pecota K V, Mwanga R O M, Catignani G L, Truong V D, Watkins R H, Ulmer M R, Yencho G C Identification of quantitative trait loci for dry-matter, starch, and β-carotene content in sweetpotato. Molecular Breeding, 28, Cervantes-Flores J C, Yencho G C, Kriegner A, Pecota K V, Faulk M A, Mwanga R O M, Sosinski B R Development of a genetic linkage map and identification of homologous linkage groups in sweetpotato using multiple-dose AFLP markers. Molecular Breeding, 21, CIP International Potato Center About sweetpotato. Li A X, Liu Q C, Wang Q M, Zhai H, Yan W Z, Zhang H Y, Li M Mapping QTLs for starch content in sweetpotato. Molecular Plant Breeding, 8, (in Chinese) Tanksley S D Mapping polygenes. Annual Review of Genetics, 27, van Ooijen J W MapQTL 4.0, software for the mapping of quantitative trait loci in experimental populations. Kyazma, Wageningen. Voorrips R E MapChart: software for the graphical presentation of linkage maps and QTLs. Journal of heredity, 93, Wang W Z, Yi F, Du S R, Wei X L, Xu L P, Cao H L Conversion table of the starch content in sweetpotato. Acta Agronomica Sinica, 15, (in Chinese) Wu J, Tan W F, He J R, Pu Z G, Wang D Y, Zhang Z S, Zhang F F, Yan W Z Construction of SRAP linkage map and QTL mapping for starch content in sweetpotato. Molecular Plant Breeding, 3, (in Chinese) Yamakawa O Developent of sweetpotato cultivars for new processing use in Japan. Tropical Agriculture, 5

6 75, Young N D QTL mapping and quantitative disease resistance in plants. Annual Review of Phytopathology, 34, Zang N, Zhai H, Gao S, Chen W, He S Z, Liu Q C Efficient production of transgenic plants using the bar gene for herbicide resistance in sweetpotato. Scientia Horticulturae, 122, (in Chinese) Zhao N Construction of high-density molecular linkage maps of sweetpotato, Ipomoea batatas (L.) Lam. Ph D thesis, China Agricultural University. (in Chinese) Table 1 QTLs detected in the F 1 population from the cross Xushu 18 Xu 781 Distance QTL Environment Linkage group Marker or from flanking marker marker LOD R 2 (%) (cm) Xu 781 DMFN_1 Langfang-2009 XU781(01.05) E31M19-21s** Langfang-2010 XU781(01.05) E31M19-21s** mean data XU781(01.05) E31M19-21s** DMFN_2 Langfang-2010 XU781(02.07) E51M28-6d Langfang-2011 XU781(02.07) E51M28-6d mean data XU781(02.07) E51M28-6d DMFN_3 Langfang-2009 XU781(03.15) E17M55-18d Langfang-2010 XU781(03.15) E17M55-18d mean data XU781(03.15) E17M55-18d DMFN_4 Langfang-2009 XU781(07.40) E27M62-2d E36M63-10ds** 4.6 Langfang-2010 XU781(07.40) E27M62-2d E36M63-10ds** 4.6 mean data XU781(07.40) E27M62-2d E36M63-10ds** 4.6 DMFN_5 Langfang-2010 XU781(08.45) E50M49-3d Langfang-2011 XU781(08.45) E50M49-3d mean data XU781(08.45) E50M49-3d DMFN_6 Langfang-2009 XU781(10.54) C61_2d Langfang-2010 XU781(10.54) C61_2d mean data XU781(10.54) C61_2d Xushu 18 DMMN_1 Langfang-2009 XUSHU18(05.27) E1M31-17ds** Langfang-2011 XUSHU18(05.27) E1M31-17ds** mean data XUSHU18(05.27) E1M31-17ds** DMMN_2 Langfang-2009 XUSHU18(06.31) E50M12-8s Langfang-2010 XUSHU18(06.31) E50M12-8s mean data XUSHU18(06.31) E50M12-8s R 2, the percentage of the variation in starch content explained by each QTL. 6

7 Fig. 1 Frequency histogram showing the distributions of starch content in the F 1 population of the cross Xushu 18 Xu 781. Black and white arrows represent the phenotypic values of each parent, respectively. Fig. 2 Mapping of QTLs for starch content on the Xu 781 linkage groups. QTLs are shown as vertical bars on the right side of the respective linkage group. QTLs with positive effect are indicated by black boxes. Markers with the same position as corresponding QTL are indicated by underlined text. 7

8 Fig. 3 Mapping of QTLs for starch content on the Xushu 18 linkage groups. QTLs are shown as vertical bars on the right side of the respective linkage group. QTLs with negative effect are indicated by shaded boxes. Markers with the same position as corresponding QTL are indicated by underlined text. 8