scholarly journals Species-Specific Detection of the Maize Pathogens Sporisorium reiliana and Ustilago maydis by Dot Blot Hybridization and PCR-Based Assays

Plant Disease ◽  
1999 ◽  
Vol 83 (4) ◽  
pp. 390-395 ◽  
Author(s):  
M. L. Xu ◽  
A. E. Melchinger ◽  
T. Lübberstedt
Keyword(s):  
Genomic Dna ◽  
Blot Hybridization ◽  
Pathogenic Fungi ◽  
Ustilago Maydis ◽  
Dot Blot ◽  
Cross Hybridization ◽  
Dot Blot Hybridization ◽  
Fungal Dna ◽  
Species Specific ◽  
Sporisorium Reiliana

Head smut of maize, caused by Sporisorium reiliana, may substantially reduce grain yield. The objective of the present study was to develop a highly specific and sensitive DNA-based assay for detection of S. reiliana and its differentiation from Ustilago maydis, a maize fungus inducing the symptomatically similar common smut disease. Plasmid libraries of S. reiliana and U. maydis were constructed using a shotgun cloning procedure. Clones containing strongly hybridizing species-specific DNA were selected by screening libraries with their own labeled genomic DNA, followed by cross-hybridization with genomic DNA of maize and other maize-pathogenic fungi. The selected clones were used to generate subclones with short insert fragments to facilitate PCR amplification for labeling and primer design for a PCR assay. Using Dig-dUTP labeled inserts, detection of less than 0.16 ng of fungal DNA was possible by dot blot hybridization. Sequences of insert fragments were determined to design primer pairs for a PCR-based assay. Primer pairs SR1 and SR3 are species-specific for S. reiliana, and UM11 is species-specific for U. maydis. The PCR-based assays can detect fungal DNA of less than 1.6 pg using SR1 and SR3, and 8 pg using UM11, irrespective of the presence of maize DNA. Use of SR1 and SR3 allowed detection of S. reiliana in the extracts of pith, node, and shank from S. reiliana-infected plants, but not in leaves. Thus, both the dot blot hybridization and the PCR-based assays provide a highly sensitive and reliable tool for detection and differentiation of corn smut caused either by S. reiliana or by U. maydis.

Species-specific DNA probes for the identification of African trypanosomes in tsetse flies

Parasitology ◽  
1988 ◽  
Vol 97 (1) ◽  
pp. 63-73 ◽  
Author(s):  
W. C. Gibson ◽  
P. Dukes ◽  
J. K. Gashumba
Keyword(s):  
Repeat Unit ◽  
Blot Hybridization ◽  
Dna Probes ◽  
Tsetse Flies ◽  
Dot Blot ◽  
Specific Group ◽  
African Trypanosomes ◽  
Dot Blot Hybridization ◽  
Species Specific

SUMMARYWe have obtained 5 specific DNA probes for African trypanosomes of the subgenera Trypanozoon and Nannomonas. Each probe consists of one repeat unit of the major repetitive DNA (satellite DNA) of each species or intra-specific group. One probe hybridized with all members of subgenus Trypanozoon (except T. equiperdum which was not tested). In subgenus Nannomonas, one probe recognized T. simiae, but 3 probes were needed to identify all stocks of T. congolense available. Each of the 3 latter probes recognized trypanosomes from one of the 3 major groups of T. congolense previously defined by isoenzyme characterization, i.e. savannah, forest and Kenya coast types. As few as 100 trypanosomes could be unequivocally identified by dot blot hybridization and individual trypanosomes could be identified by in situ hybridization. We show how this simple methodology can be used in the field for the identification of immature and mature trypanosome infections in tsetse.

Genetic homogeneity within Leishmania (L.) infantum isolated from human and dogs: the relationship with the sandfly fauna distribution in endemic areas of Nueva Esparta State, Venezuela

Parasitology ◽  
2005 ◽  
Vol 130 (6) ◽  
pp. 611-619 ◽  
Author(s):  
N. M. RODRIGUEZ ◽  
Z. DE GUGLIELMO ◽  
M. A. BARRIOS ◽  
R. M. BARRIOS ◽  
O. ZERPA ◽  
...  
Keyword(s):  
Visceral Leishmaniasis ◽  
Restriction Analysis ◽  
Blot Hybridization ◽  
Lutzomyia Longipalpis ◽  
Dot Blot ◽  
Dot Blot Hybridization ◽  
Pcr Products ◽  
Species Specific ◽  
The Relationship

Leishmania infantum has been described as a highly polymorphic group of parasites, responsible for visceral leishmaniasis and cutaneous leishmaniasis. In this paper we report the life-cycle of L. (L.) infantum in an endemic area of visceral leishmaniasis in Venezuela, by using molecular diagnosis and characterization of parasites isolated from dogs, humans with visceral leishmaniasis and sand flies. The molecular characterization was carried out by use of kDNA restriction analysis, dot-blot hybridization with species-specific probes and RFLP of the PCR products. The results demonstrated that L. (L.) infantum is the parasite responsible for VL in the island. The parasites were revealed to be genetically homogeneous with no intra-specific differences between isolates from different individuals. The highest homology of the isolates was with L. (L.) infantum from the Old World rather than with L. (L.) chagasi from the New World. Additionally, we report the geographical distribution of Lutzomyia longipalpis, and the relationship with the transmission of L. (L.) infantum in the studied area.

Detection of Human Rotaviruses in Fresh and Estuarine Waters by Dot-blot Hybridization

1991 ◽  
Vol 23 (1-3) ◽  
pp. 253-260 ◽  
Author(s):  
Abidelfatah M. Nasser ◽  
Mary K. Estes ◽  
Theodore G. Metcalf
Keyword(s):  
Fresh Water ◽  
Water Samples ◽  
Hepatitis A Virus ◽  
Blot Hybridization ◽  
Genome Segment ◽  
Detection Sensitivity ◽  
Dot Blot ◽  
Cross Hybridization ◽  
Dot Blot Hybridization ◽  
Stool Samples

This study was conducted to evaluate the suitability of dot-blot hydridization for detecting rotavirus in fresh and estuarine water samples using cloned DNA of genome segment 6 of human strain Wa and simian SA-11 rotaviruses. Probes of Wa and SA-11 cross-hybridized with heterologous RNAs of SA-11 and Wa rotaviruses under low stringency conditions. However, cross-hybridization reactions were minimized or eliminated under high stringency conditions, while homologous reactions were not affected. Wa probe did not react with representatives of the major enterovirus groups or hepatitis A virus. Hybridization detection sensitivity was not affected by organic material present in waters or proteinaceous material of eluents. Hybridization using cDNA of Wa rotavirus was found to be more sensitive than either Rotazyme ELISA or electron microscopy for detecting rotavirus in stool specimens. Dot-blot hybridization using cDNA of genome segment 6 of Wa rotavirus was found to be more sensitive than SPRIA for detecting rotavirus in estuarine and fresh water samples. Only 9 out of 54 (18.5%) estuarine and fresh water samples were positive by SPRIA, whereas dot-blot hybridization with Wa probe resulted in 21 out of 54 (38.8%) positive samples. One out of 20 (5%) stool samples and 23 out of 54 (44.6%) estuarine and fresh water samples were found to be positive by a pBR-322 plasmid probe. These results indicate that pBR-322 and related plasmids are present in high frequency in fecally polluted waters. Although different numbers of positive results were found with Wa and pBR-322 probes, more research is required to ascertain that hybridization reactions with such plasmids do not occur when using cDNA rotavirus probes.

scholarly journals Corrigendum

Parasitology ◽  
1999 ◽  
Vol 119 (6) ◽  
pp. 663-663
Keyword(s):  
Alkaline Phosphatase ◽  
Genomic Dna ◽  
Blot Hybridization ◽  
Dot Blot ◽  
Dot Blot Hybridization ◽  
Published Paper

Authors' correctionIn the recently published paper by J. Leiro et al. Parasitology119, 267–272, the legend to Fig. 4 should have read as follows:Fig. 4. Dot–blot hybridization analyses to evaluate sensitivity of the probes C38-DIG (for Microgemma caulleryi) and F9-DIG (for Tetramicra brevifilum). Lanes 1 and 3, genomic DNA of M. caulleryi and T. brevifilum respectively; lanes 2 and 4, C38 and F9 fragments in pBluescript II SK(+). Visualization was with the aid of alkaline-phosphatase-coupled anti-DIG antibody (see text).

Feasibility of Qualitative Testing of BCR-ABL and JAK2 V617F in Suspected Myeloproliperative Neoplasm (MPN) Using RT-PCR Reversed Dot Blot Hybridization (RT-PCR RDB)

2019 ◽  
Vol 19 (4) ◽  
pp. 220-227
Author(s):  
Najmiatul Masykura ◽  
Ummu Habibah ◽  
Siti Fatimah Selasih ◽  
Soegiarto Gani ◽  
Cosphiadi Irawan ◽  
...  
Keyword(s):  
Blot Hybridization ◽  
Jak2 V617f ◽  
Rt Pcr ◽  
Dot Blot ◽  
Dot Blot Hybridization

Prevalence of HPV in a Melbourne Female STD Population: Comparison of RNA and DNA Probes in Detecting HPV by Dot Blot Hybridization

1993 ◽  
Vol 4 (3) ◽  
pp. 159-164 ◽  
Author(s):  
A J Borg ◽  
G Medley ◽  
S M Garland
Keyword(s):  
Past History ◽  
Blot Hybridization ◽  
Condylomata Acuminata ◽  
Dot Blot ◽  
Hpv Dna ◽  
Sexually Transmitted ◽  
Dot Blot Hybridization ◽  
History Of ◽  
Cytological Features ◽  
Dna Types

A total of 377 women, consecutively selected as first attenders to a sexually transmitted diseases clinic in Melbourne, Australia, were examined for overt Condylomata acuminata and were screened for genital HPV DNA types 6, 11, 16, 18, 31, 33 and (35) using 2 dot blot hybridization methods. Overall, there was a 90% positivity correlation between the 2 methods with HPV DNA being detected in 12% of ectocervical samples. Overt warts were found in 15% of the women and HPV DNA was detected at the cervix in 35% with cytology predicting HPV with or without dysplasia in 27%. Thirteen percent had a past history of warts but none on examination and HPV DNA was evident in 16% while 18% had cytological features of HPV. Those with no warts evident and no past history of warts had both HPV DNA and cytological features of HPV in 7%.

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