Molecular cloning of cDNA encoding the c-kit receptor of Shiba goats and a novel alanine insertion specific to goats and sheep in the kinase insert region

The complete open reading frame (ORF) of the c-kit cDNA was cloned from a cerebellar cDNA library of the Shiba goat (Capra hircus var Shiba) with the dominant black-eyed white phenotype. The analysis of the deduced amino acid sequence revealed the presence of a single amino acid insertion (alanine) in the kinase insert (KI) region.
While the newly found alanine insertion is not correlated with the coat color phenotype of goats, it appears to be characteristic of the c-kit genes in goats and sheep. Although the biological significance of the insert remains to be investigated, its phylogenetically limited distribution will provide us with a useful and interesting tool to analyze the problems of evolution of sheep and goats in bovidae.

Cloning, sequencing and expression of stem cell factor (c-kit ligand) cDNA of brushtail possum (Trichosurus vulpecula)

By means of reverse transcription polymerase chain reaction (RT-PCR), three stem cell factor (SCF) cDNAs (822-738 bp in size) were amplified from brushtail possum ovarian poly (A)+ RNA. The largest and smallest of these cDNAs were cloned and sequenced. Characterization of these cDNAs has revealed that possum SCF has approximately 75% and 66% homology to SCF of eutherian mammals at the nucleotide level and the predicted amino acid level respectively. Nucleotide sequencing shows that the 738-bp cDNA represents an mRNA splice variant, equivalent to that found in eutherian mammals, in which an exon (84 bp) encoding a potential proteolytic cleavage site is removed. Comparison of the predicted possum SCF amino acid sequence with the predicted SCF amino acid sequences from eutherian mammals reveals conservation of all cysteine residues and 3 of 4 potential N-linked glycosylation sites.
In addition, the hydropathicity profile of the possum SCF protein is similar to that of eutherian SCF suggesting that protein conformation is conserved. Northern analysis was used to characterize possum SCF gene expression in adult ovary and testis. A major transcript of 9 kb was observed in both ovarian and testicular tissue. The conservation of the SCF gene and its predicted protein, suggests that SCF in the possum has similar biological activities to SCF in eutherian mammals.

cDNA cloning of c33-c antigen gene derived from NS3 region of Chinese HCV genome, expression in Escherichia coli and development of HCV EIA second-generation diagnostic kit.

A cDNA fragment of about 860 bp corresponding to the c33-c gene in the non-structural region 3 (NS3) of HCV genome was obtained from one plasma derived from a Chinese HCV carrier who came from Tai’ an of Shandong Province, China by the application of reverse transcription (RT) and polymerase chain reaction (PCR) techniques. After the sequence of the cDNA fragment was determined and compared with the equivalent region of the HCV-I (HCV-US) and HCV-II (HCV-BK) genomes, the nucleotide/amino acid sequence homologies were found to be 79.2%/91.3%/ and 91.3%/93.9%, respectively. The prokaryotic expression vector pBV220 was employed for the overproduction of c33-c native recombinant protein in E. coli cells.
The expression products were detected by enzyme-linked immunosorbent assay (ELISA) and Western blotting with antisera of chronic hepatitis C patients, and a molecular weight 31 kD of c33-c viral protein was shown to account for 14% of the total cellular soluble proteins. This product was extracted from the bacterial lysate by lysozyme, Triton X-100 and urea treatment, and purified through ion exchange chromatography. The purified c33-c protein combined with a branch peptide MAP-C-19 representing immunodominant epitopes on the nucleocapsid region of HCV genome was used to develop a Chinese HCV EIA 2nd-generation diagnostic kit for the detection of anti-HCV antibodies. Its specificity, sensitivity and reproducibility were all in keeping with the indexes of the national standard for the quality control of the HCV diagnostic kit. The agreement rate between our kit and Abbott company’s HCV EIA second-generation diagnostic kit was 99.33%, and the identified rate of positive anti-HCV of our kit was 2% more than that of the Abbott company’s kit.

A receptor tyrosine kinase cDNA isolated from a population of enriched primitive hematopoietic cells and exhibiting close genetic linkage to c-kit.

We have cloned a receptor tyrosine kinase cDNA, designated fetal liver kinase 1 (Flk-1), from mouse cell populations enriched for hematopoietic stem and progenitor cells. Sequence analysis of this clone reveals strong homology to the c-Kit subfamily of receptor kinases, and in particular to the Flt gene product. Chromosomal mapping shows that the Flk-1, Kit, and Pdgfra genes are closely linked. Flk-1 mRNA is expressed in primitive and more mature hematopoietic cells as well as in a wide variety of nonhematopoietic tissues.

A tool-kit for cDNA microarray and promoter analysis.

We describe two sets of programs for expediting routine tasks in analysis of cDNA microarray data and promoter sequences. The first set permits bad data points to be flagged with respect to a number of parameters and performs normalization in three different ways. It allows combining of result files into comprehensive data sets, evaluation of the quality of both technical and biological replicates and row and/or column standardization of data matrices.
The second set supports mapping ESTs in the genome, identifying the corresponding genes and recovering their promoters, analyzing promoters for transcription factor binding sites, and visual representation of the results. The programs are designed primarily for Arabidopsis thaliana researchers, but can be adapted readily for other model systems.

Novo? cDNA Kit

M1165-100 Biovision each 424.8 EUR

Novo? cDNA Kit

M1165-25 Biovision each 320.4 EUR

Evo™ cDNA Kit

M1164-100 Biovision each Ask for price

Evo™ cDNA Kit

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Novo? Transcriptome cDNA Kit

M1167-100 Biovision each 1142.4 EUR

Novo? Transcriptome cDNA Kit

M1167-25 Biovision each 529.2 EUR

Evo? cDNA Kit (gDNA Removal)

M1166-100 Biovision each 457.2 EUR

Tetro cDNA Synthesis Kit

BIO-65042 Bioline 30 Reactions Ask for price

Tetro cDNA Synthesis Kit

BIO-65043 Bioline 100 Reactions Ask for price

EVery cDNA Synthesis Kit

EVery200B-1 SBI 20 preps 307 EUR

SCRIPT cDNA Synthesis Kit

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SCRIPT cDNA Synthesis Kit

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circRNA cDNA Synthesis Kit

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SensiFAST cDNA Synthesis Kit

BIO-65053 Bioline 50 Reactions Ask for price

SensiFAST cDNA Synthesis Kit

BIO-65053/S Bioline Sample Ask for price

SensiFAST cDNA Synthesis Kit

BIO-65054 Bioline 250 Reactions Ask for price

OneScriptPlus cDNA Synthesis Kit

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OneScriptPlus cDNA Synthesis Kit

G236 ABM 100 x 20 ul reactions 202.8 EUR

Viva cDNA Synthesis Kit, 100app

cDSK01-100 Vivantis each Ask for price

Viva cDNA Synthesis Kit, 50app

cDSK01-050 Vivantis each Ask for price

OneScript Plus cDNA Synthesis Kit

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Total-Transcriptome cDNA Synthesis Kit

G904 ABM 25 reactions 268.8 EUR

Total-Transcriptome cDNA Synthesis Kit

G905 ABM 100 reactions 652.8 EUR

EpiNext Hi-Fi cDNA Synthesis Kit  

P-9004 EpiGentek
  • 381.90 EUR
  • 227.70 EUR
  • 20 Samples
  • 20 Reactions

OneScript Hot RT cDNA Synthesis Kit

E36G594 EnoGene 100 rxn 271.43 EUR

OneScript Hot RT cDNA Synthesis Kit

G594 ABM 100 rxn 295 EUR

Cloning, sequencing and expression of stem cell factor (c-kit ligand) cDNA of brushtail possum (Trichosurus vulpecula).

By means of reverse transcription polymerase chain reaction (RT-PCR), three stem cell factor (SCF) cDNAs (822-738 bp in size) were amplified from brushtail possum ovarian poly (A)+ RNA. The largest and smallest of these cDNAs were cloned and sequenced. Characterization of these cDNAs has revealed that possum SCF has approximately 75% and 66% homology to SCF of eutherian mammals at the nucleotide level and the predicted amino acid level respectively. Nucleotide sequencing shows that the 738-bp cDNA represents an mRNA splice variant, equivalent to that found in eutherian mammals, in which an exon (84 bp) encoding a potential proteolytic cleavage site is removed.
Comparison of the predicted possum SCF amino acid sequence with the predicted SCF amino acid sequences from eutherian mammals reveals conservation of all cysteine residues and 3 of 4 potential N-linked glycosylation sites. In addition, the hydropathicity profile of the possum SCF protein is similar to that of eutherian SCF suggesting that protein conformation is conserved. Northern analysis was used to characterize possum SCF gene expression in adult ovary and testis. A major transcript of 9 kb was observed in both ovarian and testicular tissue. The conservation of the SCF gene and its predicted protein, suggests that SCF in the possum has similar biological activities to SCF in eutherian mammals.

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