A FISH procedure was adopted to physical mapping rice RFLP marker RZ69 and the BAC clone 38J9 screened by RZ69 which is linked to gene Bph3 in Oryza saliva L. and O. officinalis Well ex: Watt. The FISH results showed ...A FISH procedure was adopted to physical mapping rice RFLP marker RZ69 and the BAC clone 38J9 screened by RZ69 which is linked to gene Bph3 in Oryza saliva L. and O. officinalis Well ex: Watt. The FISH results showed that both 38J9 and RZ69 were located in the middle of 4S in O. officinalis and the centromere area of 4S in O. sativa. In O. officinalis the percentage distances from the centromere to the hybridization sites were 20.00 +/- 5.40 and 22.12 +/- 3.44, the detection rates were 50.00% and 6.14%, but in O. sativa they were 0 and 0, 56.10% and 6.29% correspondingly. The results obtained from the BAC and RFLP clone were almost the same in the cultivated rice and O. officinalis. It was suggested that die marker RZ69 of the cultivated rice and its homologous sequence in O. officinalis were in the same BAC clone, the homologous sequence of Bph3 in O. officinalis should be located at the sites hybridized by both RZ69 and 38J9. Many signals on different chromosomes of O. officinalis were observed under no blocking with Cot-1 DNA, showing that the repetitive sequences were also homologous between O. sativa and O. officinalis. The identification of chromosome 4 of O. officinalis is based on comparative map with RG214 and BAC clone screened by RG214. The feasibility of comparative physical mapping performed between O. sativa and O. officinalis with rice BAC clones was discussed.展开更多
Multiple BAC-FISH is a powerful tool for modern cytogenetic researching in both animals and plants.But in cotton,this technique is unavailable due to the high percentage of repetitive sequences.
The rice BAC-DNA was used as probes and fluorescence in situ hybridization(FISH)was applied to the interphase and metaphase mitotic chromosomes of maize.To optimize the BAC-FISH technique,we respectively assayed the e...The rice BAC-DNA was used as probes and fluorescence in situ hybridization(FISH)was applied to the interphase and metaphase mitotic chromosomes of maize.To optimize the BAC-FISH technique,we respectively assayed the effect of several factors,including maize or rice genomic C_(o)t DNA used as blocking reagent of DNA,washing temperatures and FAD concentration in the washing buffer and in the hybrid solution.The results show that C_(o)t DNA of maize genome blocked the repetitive sequence of the rice BAC-DNA when the C_(o)t value was below 50.Meanwhile,it was necessary to adjust the C_(o)t value according to the different probes and their ratios.Decreasing the concentration of FAD in the hybridization mixtures,adjusting the washing rate after hybridization,and most especially,blocking the ricespecific repetitive sequences of BAC-DNA could improve the positive signals of BAC-FISH.展开更多
Oryza granulata Nees et Arn.ex Watt.is one of the three wild relatives of rice,which are the most valuable for study and utilization in China.In this study,the homol-ogy and physical locations of three rice resistance...Oryza granulata Nees et Arn.ex Watt.is one of the three wild relatives of rice,which are the most valuable for study and utilization in China.In this study,the homol-ogy and physical locations of three rice resistance genes,Glh,Bph-3 and xa-5 are comparatively analyzed between O.sativa and O.granulata by Southern blotting and fluorescence in situ hybridization(FISH).The results of Southern blotting indicate that there exist homologous sequences of the tested RFLP markers in O.granulata.By using three bacterial arti-ficial chromosome(BAC)clones scanned by the tested RFLP as probes,FISH signals are detected on both mitotic and pachytene chromosomes in O.sativa and O.granulata.Dual-color FISH demonstrates that two of the three BAC clones(14E16 and 38J9)are located on the short arm of the same chromosome pair in O.granulata.Additionally,colin-earity is shown for the two clones between O.sativa and O.granulata.Another BAC clone 44B4 is located on the end of the short arm of other chromosome pair in these two species.Although the phylogenetic relationship between O.sativa and O.granulata is the most distinct in Oryza and these two spe-cies have evidently different biological features and ecologi-cal habits,the relative lengths and arm ratios of the detected chromosomes and the relative positions of the tested clone signals on chromosomes in O.granulata are quite similar to those in O.sativa.展开更多
文摘A FISH procedure was adopted to physical mapping rice RFLP marker RZ69 and the BAC clone 38J9 screened by RZ69 which is linked to gene Bph3 in Oryza saliva L. and O. officinalis Well ex: Watt. The FISH results showed that both 38J9 and RZ69 were located in the middle of 4S in O. officinalis and the centromere area of 4S in O. sativa. In O. officinalis the percentage distances from the centromere to the hybridization sites were 20.00 +/- 5.40 and 22.12 +/- 3.44, the detection rates were 50.00% and 6.14%, but in O. sativa they were 0 and 0, 56.10% and 6.29% correspondingly. The results obtained from the BAC and RFLP clone were almost the same in the cultivated rice and O. officinalis. It was suggested that die marker RZ69 of the cultivated rice and its homologous sequence in O. officinalis were in the same BAC clone, the homologous sequence of Bph3 in O. officinalis should be located at the sites hybridized by both RZ69 and 38J9. Many signals on different chromosomes of O. officinalis were observed under no blocking with Cot-1 DNA, showing that the repetitive sequences were also homologous between O. sativa and O. officinalis. The identification of chromosome 4 of O. officinalis is based on comparative map with RG214 and BAC clone screened by RG214. The feasibility of comparative physical mapping performed between O. sativa and O. officinalis with rice BAC clones was discussed.
文摘Multiple BAC-FISH is a powerful tool for modern cytogenetic researching in both animals and plants.But in cotton,this technique is unavailable due to the high percentage of repetitive sequences.
基金This research was supported by Major State Basic Research Development Program of China(No.2001CB108806).
文摘The rice BAC-DNA was used as probes and fluorescence in situ hybridization(FISH)was applied to the interphase and metaphase mitotic chromosomes of maize.To optimize the BAC-FISH technique,we respectively assayed the effect of several factors,including maize or rice genomic C_(o)t DNA used as blocking reagent of DNA,washing temperatures and FAD concentration in the washing buffer and in the hybrid solution.The results show that C_(o)t DNA of maize genome blocked the repetitive sequence of the rice BAC-DNA when the C_(o)t value was below 50.Meanwhile,it was necessary to adjust the C_(o)t value according to the different probes and their ratios.Decreasing the concentration of FAD in the hybridization mixtures,adjusting the washing rate after hybridization,and most especially,blocking the ricespecific repetitive sequences of BAC-DNA could improve the positive signals of BAC-FISH.
基金supported by the National Natural Science Foundation of China(Grant No.39870423).
文摘Oryza granulata Nees et Arn.ex Watt.is one of the three wild relatives of rice,which are the most valuable for study and utilization in China.In this study,the homol-ogy and physical locations of three rice resistance genes,Glh,Bph-3 and xa-5 are comparatively analyzed between O.sativa and O.granulata by Southern blotting and fluorescence in situ hybridization(FISH).The results of Southern blotting indicate that there exist homologous sequences of the tested RFLP markers in O.granulata.By using three bacterial arti-ficial chromosome(BAC)clones scanned by the tested RFLP as probes,FISH signals are detected on both mitotic and pachytene chromosomes in O.sativa and O.granulata.Dual-color FISH demonstrates that two of the three BAC clones(14E16 and 38J9)are located on the short arm of the same chromosome pair in O.granulata.Additionally,colin-earity is shown for the two clones between O.sativa and O.granulata.Another BAC clone 44B4 is located on the end of the short arm of other chromosome pair in these two species.Although the phylogenetic relationship between O.sativa and O.granulata is the most distinct in Oryza and these two spe-cies have evidently different biological features and ecologi-cal habits,the relative lengths and arm ratios of the detected chromosomes and the relative positions of the tested clone signals on chromosomes in O.granulata are quite similar to those in O.sativa.
