Studies On Mapping Of Quantitative Trait Locus And Related Gene From Lycopersicon Hirsutum Affecting Fruit Characteristics

Fruit color is one of the important characters for tomato quality. Moreover, lycopene play the function on reducing the risk of certain types of cancer including carcinogenic, alimentary canal, respiratory tract and cataract formation. Therefore, to increase lycopene content has been a major objective in tomato breeding. A wide study have been conducted to clone many genes associated with the carotenoid pathway. Up to now, 108 QTLs had been mapped and near-isogenic lines had been developed. Among them, TA517 is a NIL in the Lycopersicon esculentum E6203 background and contains a 50cM introgression of chromosome 4 from L. hirsutum LA1777. As reported, several QTLs for improving many horticulturally important traits such as fruit color are involved in this segment, and GGPS gene is also localized herein.Based on previous experiments, we try to fine map QTLs and GGPS gene involved in this segment, and discuss the relationship between QTLs related fruit color and GGPS gene. The main objectives of this study were to : l)provide theory basis for manipulation of carotenoid biosynthetic pathway and perform map-based cloning of QTLs;2) obtain some tomato lines containing a shorter segment than TA517 and provide genetic basis for improving tomato qualities. Furthermore, these materials can be used for tomato breeding. The results were as follows:1. The high-density molecular genetic map for the bottom of chromosome 4 was constructed. The map contains 33 markers and the genetic distance between two markers is about 1.52cM on average. 17 new markers including 3 RAPD.2 SSR, 1 EST-BY-CLONE, 1 GGPS gene,2 COS, 1 RFLP and 7 AFLP markers were developed in this study.2. A total of 2000 F₂ individuals derived from crossing TA517 with E6203 and TA1527 with E6203 respectively were screened with the Codominant-SCAR markers T0974 and TG22 flanked TA517 , CAPS markers T1546 and E6203 flanked TA1527. This resulted in the isolation of 31 recombinant individuals that carry a shorter segment than 50cM respectively. The materials flanked by markers T0974 and RAPDM5 showed a higher lycopene content and intense fruit color, and these materials can be used for tomato breeding.3. Fine mapping of QTLs for fruit traits was performed by Single-point marker analysis. QTL qlyc that affect lycopene content was located between T0974 and RAPDM5;One QTLs cluster was identified. 12 QTLs affecting fruit color parameters and external fruit color and fruit homogenate flanked by markers TG443 and RAPDM5 were detected, and these QTLs were nominated by qwb, qwchroma, qwhue, qwA/B, qw (AIB)², qyl, qyb, qyhue, qyA/B, qy (A/B)², qya , qychroma. Two QTLs named qfw1 and qfw2 affecting fruit weight were identified and flanked by TA1546-TG443 and T0974-RAPDM5 respectively;Two QTLs qfsl,qfs2 affecting fruit shape were identified flanked by markers RAPDM5-TG22 and CT253-T1203 respectively;And one QTL qssc affecting soluble solid was detected flanked by markers T0974 and RAPDM5. QTL qlyc and QTLs involved in fruit color parameters and GGPS gene were co-localized between markers TG443 and RAPDM5. Therefore, it is hypothesized that GGPS gene must be related to the genetic regulation of carotenoid biosynthetic pathway and enhance fruit color.4. There are 5 base variations between TA517 and E6203 by analyzing partial GGPS gene sequence.12 NILs of TA517 were identified by specific primer GGPSF1R1-1F2 — GGPSF1R1-1R1. The result indicated that GGPS gene was located within the chromosomal region shared by the sub-NILsTA1467> TA1465, TA1473, TA1474, TA1527 flanked by markers T0974 and TG22. Based on analysis of new 31 recombinant individuals, GGPS gene was fine mapped further between markers P23/M59 and RAPDM5.The GGPS gene showed a significant correlation with lycopene and the chromaticity values of tomato measured with CR-400. This result indicated that base differentiations may result in different fruit color between these two materials. According to the phylogenetic tree drawn by CLUSTAL program, the GGPS obtained in this study was classified into ATGGPSl and ATGGPS3 respectively from Arabidopsis. This result suggested that the GGPS protein may supply GGPP for the biosynthesis of chlorophyll ,carotenoids , ent-kaurene and gibberellin.5. Codominant-SCAR markers were developed based on SCAR molecular marker and Multiplex-PCR. We used this new technique to identify twelve tomato NILs and screen 1000 F2 plants. The results indicated that Codominant-SCAR is characterized by good reproducibility, low cost and time consuming. It is useful for the MAS application in tomato and other crops.6. In order to develop a simple and accurate method for tomato fruit color estimation, we measured tomato fruit by using a portable Minolta chroma meter. The HUE value of tomato pericarp Jell and homogenate showed significant correlation with lycopene content ,this result suggested that lycopene content in tomato is mainly associated with flesh and jell. The HUE, A/B, and (A/B )2 color factors of homogenate produced the best regressions, these three equations produced better consistency between results from 3 regression equations and HPLC mensurements. Due to convenience and ease of using color measurements instead of long, tedious, and costly methods, this method can be used to assess the lycopene content of tomatoes, on the basis of their chromaticity values, in a very simple and easy way in any place. It also provide a possible field application for breeders.