Dot-blot hybridization was applied in this paper to detect five viruses infecting cucurbitaceous crops, Zuccini yellow mosaic virus （ZYMV）, Watermelon mosaic virus （WMV）, Cucumber mosaic virus （CMV）, Papaya ring...Dot-blot hybridization was applied in this paper to detect five viruses infecting cucurbitaceous crops, Zuccini yellow mosaic virus （ZYMV）, Watermelon mosaic virus （WMV）, Cucumber mosaic virus （CMV）, Papaya ringspot viruswatermelon strain （PRSV-W） and Squash mosaic virus （SqMV）, as a good alternative assay in seed health test and epidemiological and transgenic research. Digoxigenin-labelled cDNA probes of the five viruses were synthesized by PCR with the specific primers and applied in dot-blot hybridization to detect five viruses in crude extraction of the infected leaves. And three SqMV probes of different lengths （0.55, 1.6, and 2.7 kb, respectively） were designed to investigate the effect of hybridization. The results showed that the sensitivity for detecting the crude extraction of infected leaves by ZYMV, WMV, CMV, PRSV-W, and SqMV was down to 1：160, 1：160, 1：320, 1：160, and 1：320, respectively. Three SqMV probes of different length showed no differences on the sensitivity and specificity. The digoxigenin-labelled probes prepared by PCR could be used for accurate and rapid identification of 5 viruses infecting cucurbitaceous crops with good stabilities, sensitivities, specificity, and reproducibilifies.展开更多
A 4.34kb EcoR I fragment of kanamycin resistance plasmid from pET - 9a was purified by a DNA purification kit. The fragment was labeled with digoxigenin-dUTP with a commercial kit. A dot-blot hybridization and a colon...A 4.34kb EcoR I fragment of kanamycin resistance plasmid from pET - 9a was purified by a DNA purification kit. The fragment was labeled with digoxigenin-dUTP with a commercial kit. A dot-blot hybridization and a colony hybridization test with the probe were successfully developed for the surveillance of Kanamycin resistance to E.coli from swine. It was shown that the methods obtained 100% concordance in a positive tate. It was indicated that the method was available for the surveillance of kanamycin resistance to ? coli from swine.展开更多
A total of 44 HSV isolates from the patients was detected and typed by using HSV typespecific DNA probes labeled with Digoxigenin and 32 P respectively. 25 isolates were identified to be HSV-1, 19 were HSV-2. Compar...A total of 44 HSV isolates from the patients was detected and typed by using HSV typespecific DNA probes labeled with Digoxigenin and 32 P respectively. 25 isolates were identified to be HSV-1, 19 were HSV-2. Comparison of the typing results showed that the sensitivity and specificity of the Digoxigenin-labeled probe were equal to those of the 32 P-labeled Probe, and the typing rate of HSV isolates was 100%. The Dig-labeled probes are nonradioactive and the labeling technigue is simple, the Dig-labeled probes can be stored and reused, so they are more valuable for clinical application.展开更多
A summer-autumn (1994) molecular epidemiological study of enteric adenoviruses (EAds) in stool specimens collected in Wuhan area was conducted by using Digoxigenin-labelled DNA probes specific to EAd40, and EAd41, res...A summer-autumn (1994) molecular epidemiological study of enteric adenoviruses (EAds) in stool specimens collected in Wuhan area was conducted by using Digoxigenin-labelled DNA probes specific to EAd40, and EAd41, respectively.44 of 602 specimens were positive, among which 23 cases were identified as EAd40, 14 were EAd41, infection and 7 were dual infection. The ratio of males to females for the positive specimens was 1. 44. The infection rate of EAd40 and EAd41 each displayed no marked difference in seasons (summer and autumn) and similar age distribution was found between them. All of the two types of EAds in-fections predominated in patients with diarrhea under 3 years old- The results indicated that the Digoxigenin probe could detect DNA quantities as low as 1 pgwith satis factory specificity and the technique can be used for both clinical and ex-perimental purposes.展开更多
A new rapid technique for intrahepatic and extrahepatic HBV DNA detection by using digoxigenin labelled probe with in situ hybridization was developed.This technique has the advantage of being non-radioactive and a qu...A new rapid technique for intrahepatic and extrahepatic HBV DNA detection by using digoxigenin labelled probe with in situ hybridization was developed.This technique has the advantage of being non-radioactive and a quick procedure yielding stable results and showing a clear background.展开更多
get insight on the regulatory mechanism of Ki 67 gene expression in malignant cell cycle MethodsNon radioactive in situ hybridization (ISH) was undertaken, combined with immunohistochemistry to study the Ki 67 gene tr...get insight on the regulatory mechanism of Ki 67 gene expression in malignant cell cycle MethodsNon radioactive in situ hybridization (ISH) was undertaken, combined with immunohistochemistry to study the Ki 67 gene transcription and translation in various human cells and tissues HeLa cells and fresh colon cancer cells, tonsil, normal pancreas and pancreatic cancer tissues were used in this study A 435?bp cDNA fragment located in exon 13 of the Ki 67 antigen gene was amplified by polymerase chain reaction (PCR) Digoxigenin labelled antisense and sense RNA probes were prepared for detecting Ki 67 mRNA, combined with MIB 1 immuno^histochemistry Results Successful localization of Ki 67 mRNA in human HeLa cells, colon cancer cells, tissues specimen of the tonsil and pancreatic cancer tissue sections was accomplished by digoxigenin labelling in situ hybridization technique ISH to colon cancer cells and pancreatic cancer tissue slides showed that much stronger cytoplasm and perinuclear mRNA signals of the Ki 67 gene were present in malignant cells than in normal cells, which was in accordance with MIB 1 nuclear protein signals Conclusions A sensitive and practical in situ hybridization method for the analysis of Ki 67 antigen mRNA in human cell and tissue was developed Abnormal transcription of exon 13 of Ki 67 gene might be responsible for malignant cell proliferation in colon and pancreatic展开更多
文摘Dot-blot hybridization was applied in this paper to detect five viruses infecting cucurbitaceous crops, Zuccini yellow mosaic virus （ZYMV）, Watermelon mosaic virus （WMV）, Cucumber mosaic virus （CMV）, Papaya ringspot viruswatermelon strain （PRSV-W） and Squash mosaic virus （SqMV）, as a good alternative assay in seed health test and epidemiological and transgenic research. Digoxigenin-labelled cDNA probes of the five viruses were synthesized by PCR with the specific primers and applied in dot-blot hybridization to detect five viruses in crude extraction of the infected leaves. And three SqMV probes of different lengths （0.55, 1.6, and 2.7 kb, respectively） were designed to investigate the effect of hybridization. The results showed that the sensitivity for detecting the crude extraction of infected leaves by ZYMV, WMV, CMV, PRSV-W, and SqMV was down to 1：160, 1：160, 1：320, 1：160, and 1：320, respectively. Three SqMV probes of different length showed no differences on the sensitivity and specificity. The digoxigenin-labelled probes prepared by PCR could be used for accurate and rapid identification of 5 viruses infecting cucurbitaceous crops with good stabilities, sensitivities, specificity, and reproducibilifies.
文摘A 4.34kb EcoR I fragment of kanamycin resistance plasmid from pET - 9a was purified by a DNA purification kit. The fragment was labeled with digoxigenin-dUTP with a commercial kit. A dot-blot hybridization and a colony hybridization test with the probe were successfully developed for the surveillance of Kanamycin resistance to E.coli from swine. It was shown that the methods obtained 100% concordance in a positive tate. It was indicated that the method was available for the surveillance of kanamycin resistance to ? coli from swine.
文摘A total of 44 HSV isolates from the patients was detected and typed by using HSV typespecific DNA probes labeled with Digoxigenin and 32 P respectively. 25 isolates were identified to be HSV-1, 19 were HSV-2. Comparison of the typing results showed that the sensitivity and specificity of the Digoxigenin-labeled probe were equal to those of the 32 P-labeled Probe, and the typing rate of HSV isolates was 100%. The Dig-labeled probes are nonradioactive and the labeling technigue is simple, the Dig-labeled probes can be stored and reused, so they are more valuable for clinical application.
文摘A summer-autumn (1994) molecular epidemiological study of enteric adenoviruses (EAds) in stool specimens collected in Wuhan area was conducted by using Digoxigenin-labelled DNA probes specific to EAd40, and EAd41, respectively.44 of 602 specimens were positive, among which 23 cases were identified as EAd40, 14 were EAd41, infection and 7 were dual infection. The ratio of males to females for the positive specimens was 1. 44. The infection rate of EAd40 and EAd41 each displayed no marked difference in seasons (summer and autumn) and similar age distribution was found between them. All of the two types of EAds in-fections predominated in patients with diarrhea under 3 years old- The results indicated that the Digoxigenin probe could detect DNA quantities as low as 1 pgwith satis factory specificity and the technique can be used for both clinical and ex-perimental purposes.
文摘A new rapid technique for intrahepatic and extrahepatic HBV DNA detection by using digoxigenin labelled probe with in situ hybridization was developed.This technique has the advantage of being non-radioactive and a quick procedure yielding stable results and showing a clear background.
基金boththeNationalNaturalScienceFoundationofChina (No 396 0 0 14 1)andtheNaturalScienceFoundationofZhejiangProvince (No 396 498
文摘get insight on the regulatory mechanism of Ki 67 gene expression in malignant cell cycle MethodsNon radioactive in situ hybridization (ISH) was undertaken, combined with immunohistochemistry to study the Ki 67 gene transcription and translation in various human cells and tissues HeLa cells and fresh colon cancer cells, tonsil, normal pancreas and pancreatic cancer tissues were used in this study A 435?bp cDNA fragment located in exon 13 of the Ki 67 antigen gene was amplified by polymerase chain reaction (PCR) Digoxigenin labelled antisense and sense RNA probes were prepared for detecting Ki 67 mRNA, combined with MIB 1 immuno^histochemistry Results Successful localization of Ki 67 mRNA in human HeLa cells, colon cancer cells, tissues specimen of the tonsil and pancreatic cancer tissue sections was accomplished by digoxigenin labelling in situ hybridization technique ISH to colon cancer cells and pancreatic cancer tissue slides showed that much stronger cytoplasm and perinuclear mRNA signals of the Ki 67 gene were present in malignant cells than in normal cells, which was in accordance with MIB 1 nuclear protein signals Conclusions A sensitive and practical in situ hybridization method for the analysis of Ki 67 antigen mRNA in human cell and tissue was developed Abnormal transcription of exon 13 of Ki 67 gene might be responsible for malignant cell proliferation in colon and pancreatic
