RNA Blot Analysis for the Detection and Quantification of Plant MicroRNAs

10.3791/61394 ◽  
2020 ◽  
Author(s):  
Varsha Tirumalai ◽  
Melvin Prasad ◽  
P. V. Shivaprasad
Keyword(s):  
Blot Analysis ◽  
Detection And Quantification ◽  
Rna Blot Analysis

Multiple elements and auto-repression regulate Rox1, a repressor of hypoxic genes in Saccharomyces cerevisiae.

Genetics ◽  
1995 ◽  
Vol 139 (3) ◽  
pp. 1149-1158 ◽  
Author(s):  
J Deckert ◽  
R Perini ◽  
B Balasubramanian ◽  
R S Zitomer
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Growth ◽  
Blot Analysis ◽  
Lacz Fusion ◽  
Upstream Region ◽  
Fusion Construct ◽  
Gel Retardation ◽  
Gal1 Promoter ◽  
Rna Blot Analysis

Abstract The ROX1 gene encodes a heme-induced repressor of hypoxic genes in yeast. Using RNA blot analysis and a ROX1/lacZ fusion construct that included the ROX1 upstream region and only the first codon, we discovered that Rox1 represses its own expression. Gel-retardation experiments indicated that Rox1 was capable of binding to its own upstream region. Overexpression of Rox1 from the inducible GAL1 promoter was found to be inhibitory to cell growth. Also, we found that, as reported previously, Hap1 is partially responsible for heme-induction of ROX1, but, in addition, it also may play a role in ROX1 repression in the absence of heme. There is a second repressor of anaerobic ROX1 expression that requires the general repressor Tup1/Ssn6 for its function.

Molecular cloning of a rat uterine gap junction protein and analysis of gene expression during gestation

1991 ◽  
Vol 260 (5) ◽  
pp. E787-E793 ◽  
Author(s):  
L. M. Lang ◽  
E. C. Beyer ◽  
A. L. Schwartz ◽  
J. D. Gitlin
Keyword(s):  
Gap Junctions ◽  
Gap Junction ◽  
Cdna Clone ◽  
Blot Analysis ◽  
Nucleotide Sequence Analysis ◽  
Uterine Tissue ◽  
Coding Region ◽  
Junction Protein ◽  
Gap Junction Protein ◽  
Rna Blot Analysis

To study the molecular mechanisms controlling the rapid increase in myometrial gap junctions observed in the parturient uterus, we have isolated a full-length cDNA clone corresponding to a rat uterine gap junction protein. Nucleotide sequence analysis of the cDNA clone reveals complete identity of the coding region with that of a previously reported heart gap junction protein (connexin43). Southern blot analysis suggests that the gene encoding this gap junction protein exists as a single copy in the rat haploid genome and contains no introns within the coding region. RNA blot analysis with this gap junction cDNA reveals a single 3.0-kb mRNA in uterine tissue without changes in transcript size throughout gestation. When normalized to the amount of 28S rRNA, the relative abundance of the connexin43 transcript in uterine tissue is quite constant between the nonpregnant state, during gestation, intrapartum, and postpartum. Similar size transcripts are shown by RNA blot analysis to be present in heart, lung, liver, brain, and skeletal muscle, and these transcripts are identified by the same 3'-nontranslated sequence probe. The results of these studies suggest that rat connexin43 is encoded by a single gene that is transcribed to identical transcripts in heart, uterus, and other tissues. They further suggest that changes in the abundance of connexin43 transcript are unlikely to be responsible for the abrupt increase in connexin43-containing myometrial gap junctions at term.

A procedure for the small-scale isolation of plant RNA suitable for RNA blot analysis

1988 ◽  
Vol 172 (1) ◽  
pp. 279-283 ◽  
Author(s):  
Gregory J. Wadsworth ◽  
Margaret G. Redinbaugh ◽  
John G. Scandalios
Keyword(s):  
Blot Analysis ◽  
Small Scale ◽  
Rna Blot Analysis

scholarly journals First Report of Leek yellow stripe virus in Garlic in the State of Guanajuato, Mexico

Plant Disease ◽  
2006 ◽  
Vol 90 (11) ◽  
pp. 1458-1458 ◽  
Author(s):  
L. Pérez-Moreno ◽  
Z. Córdova-Rosales ◽  
E. Barboza-Corona ◽  
Rafael Ramírez-Malagón ◽  
J. Ramírez-Lúa ◽  
...  
Keyword(s):  
Coat Protein ◽  
Blot Analysis ◽  
The State ◽  
Latent Virus ◽  
Enzyme Linked Immunosorbent Assay ◽  
First Report ◽  
Leek Yellow Stripe Virus ◽  
Virus Complex ◽  
Yellow Stripe ◽  
Rna Blot Analysis

Garlic (Allium sativum L.) can be affected by a virus complex (1) consisting of two potyviruses, Onion yellow dwarf virus (OYDV) and Leek yellow stripe virus (LYSV), and two carlaviruses, Garlic common latent virus (GCLV) and Shallot latent virus (SLV) (1). To identify the components of the virus complex that could be present in garlic plants in Guanajuato State, which is the second largest garlic producer in the country and where presumptive viral symptoms were initially observed in December 2004, a survey was carried out in six locations: San Miguel de Allende and San Luis de la Paz in northern Guanajuato; Irapuato and Villagrán in the central region; and Salamanca and Valle de Santiago in the southern part of the state. Enzyme-linked immunosorbent assay (ELISA) was carried out to detect LYSV, OYDV, GCLV, and SLV in 195 garlic leaf samples collected during January 2005 from plants with leaf yellow stripe, mosaic, enation, deformation, or dwarfism symptoms. A set of primers, previously reported and specific to the coat protein cistron of LYSV (1), were synthesized and used in a reverse transcriptase-polymerase chain reaction (RT-PCR). The amplified product (1,020 nucleotides) was cloned into plasmid pGEM T-Easy (Promega, Madison, WI) and sequenced (Gen-bank Accession No. DQ841554). Sequence analysis showed that the cloned DNA fragment shared 97% similarity with the coat protein cistron of LYSV isolate no. 3 from Okinawa (GenBank Accession No. AB194632). The fragment was then radioactively labelled and used as a probe in the RNA blot analysis of all samples to confirm the ELISA results of LYSV. Of the 195 samples, 64 tested positive by RNA blot analysis. Forty-one of these were also positive by ELISA for LYSV. Preliminary, positive ELISA results were also obtained for OYDV, GCLV and SLV. To our knowledge, this is the first report of LYSV in the State of Guanajuato and in Mexico. The correct identification of viruses present in garlic will help to use the appropriate strategies to reduce viral incidence in this garlic-producing region. Reference: (1) T. V. M. Fajardo et al. Fitopatol. Bras. 26:619, 2001.

Northern blot analysis for detection and quantification of RNA in pancreatic cancer cells and tissues

2008 ◽  
Vol 4 (1) ◽  
pp. 37-43 ◽  
Author(s):  
Sylvia Streit ◽  
Christoph W Michalski ◽  
Mert Erkan ◽  
Jörg Kleeff ◽  
Helmut Friess
Keyword(s):  
Pancreatic Cancer ◽  
Cancer Cells ◽  
Northern Blot ◽  
Blot Analysis ◽  
Northern Blot Analysis ◽  
Pancreatic Cancer Cells ◽  
Detection And Quantification

scholarly journals Developmental regulation of calmodulin gene expression in rat brain and skeletal muscle.

1991 ◽  
Vol 2 (10) ◽  
pp. 819-826 ◽  
Author(s):  
J Weinman ◽  
B Della Gaspera ◽  
A Dautigny ◽  
D Pham Dinh ◽  
J Wang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Skeletal Muscle ◽  
Rat Brain ◽  
Blot Analysis ◽  
Temporal Pattern ◽  
Developmental Regulation ◽  
Mrna Levels ◽  
Calmodulin Gene ◽  
The Brain ◽  
Rna Blot Analysis

Three different calmodulin genes that encode the identical protein have been identified in the rat (Nojima, 1989); however, calmodulin gene expression at the various stages of tissue differentiation and maturation has not been previously determined. We have quantitated the content of mRNAs encoding calmodulin in the developing brain and skeletal muscle using RNA blot analysis with three specific cDNA probes. Our results show that five species of calmodulin mRNAs: 4.0 and 1.7 kb for CaM I, 1.4 kb for CaM II, and 2.3 and 0.8 kb for CaM III are detectable at all ages in the brain as well as in skeletal muscle but exhibit a tissue-specific developmental pattern of expression. The comparison of the temporal pattern of calmodulin gene expression with both mitotic activity, as demonstrated by cyclin A mRNA levels, and differentiation and maturation of specific brain or muscle regions is consistent with calmodulin involvement in development.

Rainbow Trout (Salmo gairdneri) Growth Hormone: in vitro Translation of Pituitary RNA and Product Analysis

1986 ◽  
Vol 43 (7) ◽  
pp. 1327-1331 ◽  
Author(s):  
L. B. Agellon ◽  
T. T. Chen ◽  
R. J. Van Beneden ◽  
R. A. Sonstegard ◽  
G. F. Wagner ◽  
...  
Keyword(s):  
Growth Hormone ◽  
Rainbow Trout ◽  
Blot Analysis ◽  
Immunoblot Analysis ◽  
In Vitro Translation ◽  
Salmo Gairdneri ◽  
Western Immunoblot ◽  
Western Immunoblot Analysis ◽  
Rna Blot Analysis

We have used RNA blot analysis, in vitro translation, and protein immunoblot analysis to characterize mRNA in the pituitary glands of rapidly growing rainbow trout (Salmo gairdneri). Cell-free translation products of total RNA, analyzed by direct immunoprecipitation or western immunoblot analysis with an antiserum to chum salmon (Oncorphynchus keta) growth hormone (GH), indicate that rainbow trout pregrowth hormone (pre-GH) is a polypeptide of 25 000 Da. RNA blot analysis of total pituitary RNA to its cDNA, using total liver RNA as a competitor, revealed the presence of at least four size classes of pituitary mRNA sequences. One class, with a size smaller than 18S rRNA, is the predominant mRNA component. By in vitro translation and western immunoblot analysis, the pre-GH mRNA activity is associated with this class of mRNA sequences. These results suggest that rainbow trout pre-GH mRNA is in the same size range as that of mammalian GH mRNA.

scholarly journals 058 Cloning of a Defensin-related Gene and Its Expression during Dormancy and Fruit Development in Peach [Prunus persica (L.) Batsch]

HortScience ◽  
1999 ◽  
Vol 34 (3) ◽  
pp. 451B-451
Author(s):  
M. Wisniewski ◽  
T. Artlip ◽  
R. Webb ◽  
C. Bassett ◽  
A. Callahan
Keyword(s):  
Protease Inhibitor ◽  
Fruit Development ◽  
Blot Analysis ◽  
Prunus Persica ◽  
Extracellular Proteins ◽  
Full Length ◽  
Amino Terminal ◽  
Partial Clone ◽  
Extracellular Transport ◽  
Rna Blot Analysis

During the past several years we have been involved in identifying seasonally regulated proteins and genes from peach bark. In the present study, we describe the cloning of a protease inhibitor from a cDNA library made from winter bark tissues. A partial clone obtained from the library was extended to full length by 5' RACE. The full-length cDNA clone (final3b) is 613 bp in length, not including the poly A+ tail. The open reading frame of 237 bp codes for a 79 amino acid protease inhibitor related to the defensin family of proteins. This family of small, cysteine-rich, extracellular proteins play a role in the plantís defense response through their antifungal properties. Sequence comparison of the encoded protein using BLAST analysis revealed significant homology to protease inhibitors from Glycine max, Arabidopsis thaliana, and a defensin protein from bell pepper (Capsicum annuum). Similar to these other cysteine-rich proteins, the peach defensin contains a consensus cys arrangement and is predicted to have an amino terminal signal peptide, presumably targeting it for extracellular transport. RNA-blot analysis indicated that the gene is seasonally expressed in bark tissues of 1-year-old shoots. Transcript abundance of final3b increased in the fall, reached a peak in midwinter and then decreased. The gene was also expressed during early stages of fruit development. RNA-blot analysis of the gene in other tissues, and in response to environmental stress and wounding, is in progress.

Protein gel blot analysis

2007 ◽  
Author(s):  
Dominique Loqué ◽  
Wolf Frommer
Keyword(s):  
Blot Analysis ◽  
Protein Gel
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