Isolation of epiblast-specific cDNA clones by differential hybridization with polymerase chain reaction-amplified probes derived from single embryos

1992 ◽  
Vol 32 (4) ◽  
pp. 339-348 ◽  
Author(s):  
Susannah Varmuza ◽  
Peri Tate
Keyword(s):  
Polymerase Chain Reaction ◽  
Cdna Clones ◽  
Chain Reaction ◽  
Differential Hybridization ◽  
Polymerase Chain

scholarly journals The δ-subunit of ATP synthase from bovine heart mitochondria. Complementary DNA sequence of its import precursor cloned with the aid of the polymerase chain reaction

1990 ◽  
Vol 266 (2) ◽  
pp. 421-426 ◽  
Author(s):  
M J Runswick ◽  
S M Medd ◽  
J E Walker
Keyword(s):  
Polymerase Chain Reaction ◽  
Atp Synthase ◽  
Protein Sequence ◽  
Heart Mitochondria ◽  
Cdna Clones ◽  
Mature Protein ◽  
Chain Reaction ◽  
Bovine Heart ◽  
Polymerase Chain ◽  
Delta Subunit

The delta-subunit of ATP synthase from bovine heart mitochondria is part of the extrinsic membrane domain, F1-ATPase. The mature protein is 146 amino acids in length and its function is obscure. It is encoded by a nuclear gene and is imported into the organelle. Two mixtures of oligonucleotides 17 bases long, designed on the basis of the known protein sequence, have been synthesized and employed as primers on bovine cDNA in the polymerase chain reaction. By this means a segment of bovine cDNA encoding part of the delta-subunit has been amplified, and this DNA segment has been employed to identify related cDNA clones in a library. These clones encode the mitochondrial import precursor of the delta-subunit; the protein sequence of the mature protein deduced from it is exactly the same as that determined earlier by direct sequence analysis. The clones have also been used to show that both the bovine and human genomes seem to contain a single gene for the delta-subunit.

scholarly journals Sheep 6-phosphogluconate dehydrogenase. Revised protein sequence based upon the sequences of cDNA clones obtained with the polymerase chain reaction

1992 ◽  
Vol 288 (3) ◽  
pp. 1061-1067 ◽  
Author(s):  
D O Somers ◽  
S M Medd ◽  
J E Walker ◽  
M J Adams
Keyword(s):  
Polymerase Chain Reaction ◽  
Protein Sequence ◽  
Direct Analysis ◽  
Pentose Phosphate ◽  
Cdna Clones ◽  
Mature Protein ◽  
Chain Reaction ◽  
Phosphogluconate Dehydrogenase ◽  
Sheep Liver ◽  
Polymerase Chain

Sheep liver 6-phosphogluconate dehydrogenase (6-PGDH) is an enzyme of the pentose phosphate pathway. Evidence has appeared which suggests that the 6-PGDH protein sequence determined previously by direct analysis of the protein isolated from ovine liver is incorrect. Determining the enzyme's DNA sequence was considered to be the best way of solving the problem. In the first instance, a degenerate forward and a degenerate reverse primer were designed on the basis of the known protein sequence, and a partial-length cDNA clone was isolated from total sheep liver cDNA using the polymerase chain reaction. The clone encoded the expected part of the protein sequence. The clone was unsuccessfully used as a prime-cut probe to screen a sheep liver library and a bovine heart library. As a result, the polymerase chain reaction was utilized again to successfully generate a family of overlapping cDNA clones encoding a mature protein of 482 amino acids. The mature protein sequence encoded by the cDNA differs significantly from the sequence derived by direct analysis of the protein, but on closer examination the fundamental difference is caused by the incorrect placement of three enzyme fragments obtained by cyanogen bromide cleavage during the direct sequence analysis of the protein. Placing the fragments in the correct order results in the two sequences being virtually identical except for some minor amino acid changes between the amide and acid forms, and a small number of deletions and insertions.

scholarly journals Identification of novel protein tyrosine phosphatases of hematopoietic cells by polymerase chain reaction amplification

Blood ◽  
1991 ◽  
Vol 78 (9) ◽  
pp. 2222-2228 ◽  
Author(s):  
T Yi ◽  
JL Cleveland ◽  
JN Ihle
Keyword(s):  
Polymerase Chain Reaction ◽  
Polymerase Chain Reaction Amplification ◽  
Hematopoietic Cells ◽  
Protein Tyrosine Phosphatases ◽  
Cdna Clones ◽  
Tyrosine Phosphatases ◽  
Chain Reaction ◽  
Protein Tyrosine ◽  
Cdna Sequences ◽  
Polymerase Chain

Polymerase chain reaction (PCR) conditions were used to amplify cDNAs that encode putative protein tyrosine phosphatases (PTPs) from a murine interleukin-3-dependent myeloid cell line. Primers for the reactions were based on conserved sequences of the catalytic domain that are shared among all known PTPs. Sequencing of 100 PCR-amplified cDNA clones identified seven different cDNA sequences. Two of these sequences were identical to the murine PTP genes Ly5/CD45/LCA and LRP/R- PTP-alpha. Two of the cDNA sequences were 95% identical to human PTP epsilon (HPTP epsilon) and rat brain PTP (PTP1B), respectively, and are likely to represent their murine homologs. Three of the cDNA sequences encoded novel potential PTPs. One of the putative PTPs was ubiquitously expressed while a second was predominantly expressed in brain, kidney, and liver and at much lower levels in a variety of other cell tkpes and tissues. The third novel putative phosphatase was expressed primarily in hematopoietic cells and tissues in a pattern that was comparable with Ly5/CD45/LCA. Further characterization of these novel PTPs will provide insights into the growth regulation of hematopoietic cells.

scholarly journals Induction of a Grapevine Germin-Like Protein (VvGLP3) Gene Is Closely Linked to the Site of Erysiphe necator Infection: A Possible Role in Defense?

2007 ◽  
Vol 20 (9) ◽  
pp. 1112-1125 ◽  
Author(s):  
Dale Godfrey ◽  
Amanda J. Able ◽  
Ian B. Dry
Keyword(s):  
Polymerase Chain Reaction ◽  
Cell Wall ◽  
Recombinant Protein ◽  
Plasmopara Viticola ◽  
Cdna Clones ◽  
Chain Reaction ◽  
Erysiphe Necator ◽  
Fusion Construct ◽  
Polymerase Chain ◽  
Insight Into

Germin-like proteins (GLP) have various proposed roles in plant development and defense. Seven novel GLP cDNA clones were isolated from grapevine (Vitis vinifera cv. Chardonnay). Reverse transcriptase-polymerase chain reaction expression analysis revealed that the VvGLP genes exhibit diverse and highly specific patterns of expression in response to a variety of abiotic and biotic treatments, including challenge by Erysiphe necator, Plasmopara viticola, and Botrytis cinerea, suggesting a diversity of roles for each of the GLP family members. Significantly, one of the grapevine GLP genes, VvGLP3, is induced specifically by E. necator infection and expression is closely linked to the site of infection. Subcellular localization of VvGLP3 determined by transient expression of a VvGLP3:GFP fusion construct in onion cells indicated that the recombinant protein was targeted to the cell wall. Recombinant VvGLP3 was successfully expressed in Arabidopsis thaliana and the partially purified recombinant protein was demonstrated to have superoxide dismutase activity. This data has provided an insight into the diverse nature of the GLP family in grapevine and suggests that VvGLP3 may be involved in the defense response against E. necator.

scholarly journals Identification of novel protein tyrosine phosphatases of hematopoietic cells by polymerase chain reaction amplification

Blood ◽  
1991 ◽  
Vol 78 (9) ◽  
pp. 2222-2228 ◽  
Author(s):  
T Yi ◽  
JL Cleveland ◽  
JN Ihle
Keyword(s):  
Polymerase Chain Reaction ◽  
Polymerase Chain Reaction Amplification ◽  
Hematopoietic Cells ◽  
Protein Tyrosine Phosphatases ◽  
Cdna Clones ◽  
Tyrosine Phosphatases ◽  
Chain Reaction ◽  
Protein Tyrosine ◽  
Cdna Sequences ◽  
Polymerase Chain

Abstract Polymerase chain reaction (PCR) conditions were used to amplify cDNAs that encode putative protein tyrosine phosphatases (PTPs) from a murine interleukin-3-dependent myeloid cell line. Primers for the reactions were based on conserved sequences of the catalytic domain that are shared among all known PTPs. Sequencing of 100 PCR-amplified cDNA clones identified seven different cDNA sequences. Two of these sequences were identical to the murine PTP genes Ly5/CD45/LCA and LRP/R- PTP-alpha. Two of the cDNA sequences were 95% identical to human PTP epsilon (HPTP epsilon) and rat brain PTP (PTP1B), respectively, and are likely to represent their murine homologs. Three of the cDNA sequences encoded novel potential PTPs. One of the putative PTPs was ubiquitously expressed while a second was predominantly expressed in brain, kidney, and liver and at much lower levels in a variety of other cell tkpes and tissues. The third novel putative phosphatase was expressed primarily in hematopoietic cells and tissues in a pattern that was comparable with Ly5/CD45/LCA. Further characterization of these novel PTPs will provide insights into the growth regulation of hematopoietic cells.

High performance Nanogold™-in situ hybridzation and its use in the detection of hybridized and PCR-amplified microscopical preparations

1996 ◽  
Vol 54 ◽  
pp. 896-897
Author(s):  
G. W. Hacker ◽  
I. Zehbe ◽  
J. Hainfeld ◽  
A.-H. Graf ◽  
C. Hauser-Kronberger ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Messenger Rna ◽  
High Performance ◽  
Detection System ◽  
Direct Methods ◽  
Genetic Alterations ◽  
Chain Reaction ◽  
Protein Encoding ◽  
Polymerase Chain

In situ hybridization (ISH) with biotin-labeled probes is increasingly used in histology, histopathology and molecular biology, to detect genetic nucleic acid sequences of interest, such as viruses, genetic alterations and peptide-/protein-encoding messenger RNA (mRNA). In situ polymerase chain reaction (PCR) (PCR in situ hybridization = PISH) and the new in situ self-sustained sequence replication-based amplification (3SR) method even allow the detection of single copies of DNA or RNA in cytological and histological material. However, there is a number of considerable problems with the in situ PCR methods available today: False positives due to mis-priming of DNA breakdown products contained in several types of cells causing non-specific incorporation of label in direct methods, and re-diffusion artefacts of amplicons into previously negative cells have been observed. To avoid these problems, super-sensitive ISH procedures can be used, and it is well known that the sensitivity and outcome of these methods partially depend on the detection system used.

1504: Molecular Diagnosis and Staging of Prostate Cancer Using Clusterin in Real Time Reverse Transcription Polymerase Chain Reaction (RT-PCR)

2006 ◽  
Vol 175 (4S) ◽  
pp. 485-486
Author(s):  
Sabarinath B. Nair ◽  
Christodoulos Pipinikas ◽  
Roger Kirby ◽  
Nick Carter ◽  
Christiane Fenske
Keyword(s):  
Prostate Cancer ◽  
Polymerase Chain Reaction ◽  
Real Time ◽  
Molecular Diagnosis ◽  
Reverse Transcription ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain
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