莲藕组织富含多糖、脂质、酚类等物质,用一般的方法较难提取高质量的RNA。在改进前人方法的基础上,建立了一种高效、简单的CTAB-L iC l提取法,能快速提取高质量的莲藕组织总RNA,并且产率高、完整性好、纯度高,能进一步满足RT-PCR等分子...莲藕组织富含多糖、脂质、酚类等物质,用一般的方法较难提取高质量的RNA。在改进前人方法的基础上,建立了一种高效、简单的CTAB-L iC l提取法,能快速提取高质量的莲藕组织总RNA,并且产率高、完整性好、纯度高,能进一步满足RT-PCR等分子生物学实验的需要。此外,该方法也适用于其它富含多糖、脂质、酚类等物质的植物组织总RNA的提取。展开更多
The techniques of in situ hybridization (ISH) are widely adopted for analyzing the genetic make_up and RNA expression patterns of individual cells. There are four main criterions for evaluating this technique, includi...The techniques of in situ hybridization (ISH) are widely adopted for analyzing the genetic make_up and RNA expression patterns of individual cells. There are four main criterions for evaluating this technique, including detection sensitivity, resolution, capacity and specificity. This review focuses on a number of advances made over the last years in the fluorescence in situ hybridization (FISH). These advances can be catagorized into several branches as follows: (1) Multicolor_FISH (mFISH), including conventional mFISH, combinatorial FISH, ratio labelling FISH, multicolor chromosome painting and comparative genomic hybridization (CGH); (2) Extended DNA fiber_FISH (EDF_FISH), including quantitative DNA fiber mapping (QDFM), molecular combing (MC) and dynamic molecular combing (DMC); (3) In situ PCR_based FISH; (4) Bacterial (or yeast) artificial chromosome_FISH (BAC_FISH or YAC_FISH); (5) Tyramide signal amplification_FISH (TSA_FISH); (6) Polypeptide nucleic acid_FISH (PNA_FISH) and (7) padlock_FISH.展开更多
The distribution of repetitive DNAs along chromosomes is one of the crucial elements for understanding the organization and the evolution of plant genomes. Using a modified genomic in situ hybridization （GISH） proce...The distribution of repetitive DNAs along chromosomes is one of the crucial elements for understanding the organization and the evolution of plant genomes. Using a modified genomic in situ hybridization （GISH） procedure, fluorescence in situ hybridization （FISH） with genomic DNA to their own chromosomes （called self-genomic in situ hybridization, self-GISH） was carried out in six selected plant species with different genome size and amount of repetitive DNA. Nonuniform distribution of the fluorescent labeled probe DNA was observed on the chromosomes of all the species that were tested. The signal patterns varied among species and were related to the genome size. The chromosomes of the small Arabidopsis genome were labeled almost only in the pericentromeric regions and the nucleolus organizer regions （NORs）. The signals in the relatively small genomes, rice, sorghum, and Brassica oleracea var. capitata L., were dispersed along the chromosome lengths, with a predominant distribution in the pericentromeric or proximal regions and some heterochromatic arms. All chromosomes of the large genomes, maize and barley, were densely labeled with strongly labeled regions and weakly labeled or unlabeled regions being arranged alternatively throughout the lengths. In addition, enhanced signal bands were shown in all pericentromeres and the NORs in B. oleracea var. capitata, and in all pericentromeric regions and certain intercalary sites in barley. The enhanced signal band pattern in barley was found consistent with the N-banding pattern of this species. The GISH with self-genomic DNA was compared with FISH with Cot-1 DNA in rice, and their signal patterns are found to be basically consistent. Our results showed that the self-GISH signals actually reflected the hybridization of genomic repetitive DNAs to the chromosomes, thus the self-GISH technique would be useful for revealing the distribution of the regions where repetitive DNAs concentrate along chromosomes and some chromatin differentiation associated with repetitive DNAs in plants.展开更多
文摘莲藕组织富含多糖、脂质、酚类等物质,用一般的方法较难提取高质量的RNA。在改进前人方法的基础上,建立了一种高效、简单的CTAB-L iC l提取法,能快速提取高质量的莲藕组织总RNA,并且产率高、完整性好、纯度高,能进一步满足RT-PCR等分子生物学实验的需要。此外,该方法也适用于其它富含多糖、脂质、酚类等物质的植物组织总RNA的提取。
文摘The techniques of in situ hybridization (ISH) are widely adopted for analyzing the genetic make_up and RNA expression patterns of individual cells. There are four main criterions for evaluating this technique, including detection sensitivity, resolution, capacity and specificity. This review focuses on a number of advances made over the last years in the fluorescence in situ hybridization (FISH). These advances can be catagorized into several branches as follows: (1) Multicolor_FISH (mFISH), including conventional mFISH, combinatorial FISH, ratio labelling FISH, multicolor chromosome painting and comparative genomic hybridization (CGH); (2) Extended DNA fiber_FISH (EDF_FISH), including quantitative DNA fiber mapping (QDFM), molecular combing (MC) and dynamic molecular combing (DMC); (3) In situ PCR_based FISH; (4) Bacterial (or yeast) artificial chromosome_FISH (BAC_FISH or YAC_FISH); (5) Tyramide signal amplification_FISH (TSA_FISH); (6) Polypeptide nucleic acid_FISH (PNA_FISH) and (7) padlock_FISH.
基金This work was supported by the National Natural Sciences Foundation of China (No. 39870423).
文摘The distribution of repetitive DNAs along chromosomes is one of the crucial elements for understanding the organization and the evolution of plant genomes. Using a modified genomic in situ hybridization （GISH） procedure, fluorescence in situ hybridization （FISH） with genomic DNA to their own chromosomes （called self-genomic in situ hybridization, self-GISH） was carried out in six selected plant species with different genome size and amount of repetitive DNA. Nonuniform distribution of the fluorescent labeled probe DNA was observed on the chromosomes of all the species that were tested. The signal patterns varied among species and were related to the genome size. The chromosomes of the small Arabidopsis genome were labeled almost only in the pericentromeric regions and the nucleolus organizer regions （NORs）. The signals in the relatively small genomes, rice, sorghum, and Brassica oleracea var. capitata L., were dispersed along the chromosome lengths, with a predominant distribution in the pericentromeric or proximal regions and some heterochromatic arms. All chromosomes of the large genomes, maize and barley, were densely labeled with strongly labeled regions and weakly labeled or unlabeled regions being arranged alternatively throughout the lengths. In addition, enhanced signal bands were shown in all pericentromeres and the NORs in B. oleracea var. capitata, and in all pericentromeric regions and certain intercalary sites in barley. The enhanced signal band pattern in barley was found consistent with the N-banding pattern of this species. The GISH with self-genomic DNA was compared with FISH with Cot-1 DNA in rice, and their signal patterns are found to be basically consistent. Our results showed that the self-GISH signals actually reflected the hybridization of genomic repetitive DNAs to the chromosomes, thus the self-GISH technique would be useful for revealing the distribution of the regions where repetitive DNAs concentrate along chromosomes and some chromatin differentiation associated with repetitive DNAs in plants.
