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Biomedical subjects

K A Marcker

Publications and source records attributed to K A Marcker.

At least 19 recordsLinked to original sources
PubMed1992

Isolation of DNA markers linked to a beet cyst nematode resistance locus in Beta patellaris and Beta procumbens.

In cultivated beet no useful level of resistance of the beet cyst nematode (BCN) Heterodera schachtii Schm. has been found, unlike the situation in wild species of the section Procumbentes. Stable introgression of resistance genes from the wild species into Beta vulgaris has not been achieved, but resistant monosomic additions (2n = 18 + 1), diploids of B. vulgaris with an extra alien chromosome carrying the resistance locus, have been obtained. Here we describe a new series of resistant monosomic fragment addition material of B. patellaris chromosome 1 (pat-1). We further describe the cloning of a single-copy DNA marker that specifically hybridizes with a monosomic addition fragment of approximately 8 Mb (AN5-90) carrying the BCN resistance locus. This marker and another fragment-specific, single-copy DNA marker probably flank the BCN locus on the addition fragment present in the AN5-203 material, which is approximately 19 Mb in size. Furthermore, several specific repetitive DNA markers have been isolated, one of which hybridizes to AN5-90 and also to DNA from a smaller DNA segment of Beta procumbens, present in line B883, carrying a BCN resistance locus introgressed into the B. vulgaris genome. This suggests that the specific repetitive marker is closely linked to the BCN locus.

Animals↗
PubMed1991

Characterization and genomic organization of a highly expressed late nodulin gene subfamily in soybeans.

A soybean nodulin cDNA clone (E41) hybrid-selected mRNA for three in vitro translation products with apparent molecular weights of 26 kDa, 25 kDa and 24 kDa. Based on Southern analysis of soybean genomic DNA, combined with mapping and sequencing of genomic clones, we identified four genes that are related to E41, one of which was identified to be the previously characterized N-20 gene. Our data indicate the linkage of three of the genes, of which one is a truncated version and suggest that they originated by gene duplication combined with deletion and conversion. The genes are highly expressed and we postulate that the sequence conservation in the 5' and 3' flanking regions of all four genes, has a functional role in their expression. Hybrid-selected translation products of E41 are not immunoprecipitable with antibody to the soluble fraction of nodules suggesting that they are membrane associated. The N-20 gene, which is a member of this gene subfamily, showed sequence similarity to four previously characterized nodulin genes and a phylogenetic tree is proposed based on the extent of sequence similarity.

Base Sequence↗
PubMed1991

A two-component nodule-specific enhancer in the soybean N23 gene promoter.

The two positive cis elements in the soybean nodulin N23 gene promoter were investigated in transgenic Lotus corniculatus plants and shown to constitute a two-component nodule-specific enhancer. Equal quantitative contributions from the two components were suggested by the similar expression level of chimeric N23-chloramphenicol acetyltransferase genes after deletion of either the distal positive element (PE-A, -320 to -298) or the proximal positive element (PE-B, -257 to -165). A combined effect of the two elements was indicated by orientation-dependent effects in the N23 promoter, and by the observation that neither PE-A nor PE-B separately was able to confer any activity to the cauliflower mosaic virus 35S minimal promoter. Reactivation of the minimal N23 and the minimal cauliflower mosaic virus 35S promoters by the inverted complete element (PE-AB) further suggested that PE-AB is a nodule-specific enhancer containing two equally strong enhancer components. Two 12-bp sequence motifs, InvA and InvB, constituting an inverted repeat, were identified as the core of the enhancer components PE-A and PE-B, respectively. Point mutations in InvA or InvB resulted in lower expression levels and mutations in both abolished enhancer activity. Point mutations in two nodulin consensus sequences, 5'-CTCTT and 5'-AAAGAT located downstream of PE-AB, resulted in a decreased level of expression, confirming the involvement of these two motifs in nodulin gene expression. The binding site for the nodule-specific trans-acting factor, NAT2, present in the PE-A segment, was removed without affecting expression significantly. This interaction is, therefore, dispensable for enhancer activity.

Base Sequence↗
PubMed1990

Interdependence and nodule specificity of cis-acting regulatory elements in the soybean leghemoglobin lbc3 and N23 gene promoters.

The qualitative and quantitative contributions of four separate cis-acting DNA elements controlling the root nodule-specific soybean leghemoglobin lbc3 gene were analyzed in transgenic Lotus corniculatus plants. Expression from internal deletions in the 5' region between positions -49 and -1956 was monitored from a CAT reporter gene. The strong positive element (SPE; -1090, -947) responsible for high-level expression was demonstrated to be an organ-specific element by deleting proximal nodule-specific control elements. Deletion of the downstream qualitative organ-specific element (OSE; -139, -102) containing the putative nodulin consensus sequences 5'AAAGAT and 5'CTCTT resulted in a low expression level. Efficient SPE enhancement is therefore dependent on the organ-specific element, which by itself does not enhance expression. This quantitative effect of the immediate upstream region carrying the consensus sequences was also found in hybrid promoter studies using the soybean nodulin N23 gene promoter, suggesting the involvement of these motifs in a regulatory mechanism for nodulin genes. Deletion of the lbc3 negative element (NE, -102, -49) linking the SPE and OSE onto the TATA box did not lead to unregulated expression. These results indicate that interaction between positive, negative and neutral qualitative elements controls lbc3 expression. Binding of the nuclear protein NAT2 at the lbc3 weak positive element (WPE; -230, -170) is probably not directly required for this mechanism.

Base Sequence↗
PubMed1990

HMG I-like proteins from leaf and nodule nuclei interact with different AT motifs in soybean nodulin promoters.

Three different nuclear factors recognizing short AT-rich DNA sequences were identified in different organs of soybean. One factor (NAT2) was found to be present in mature nodules, another factor (NAT1) was detected in roots and nodules, and a third one (LAT1) was only observed in leaves. All three factors recognized several DNA sequences in the promoter region of the soybean nodulin N23 gene. Footprinting, deletion, and point mutation analyses revealed different binding properties for all three factors and further showed that even single base pair substitutions had a dramatic effect on binding affinity. The LAT1 and NAT1 factors were released from chromatin by extraction with a low-salt buffer and were soluble in 2% trichloroacetic acid, implying a relationship to high-mobility group (HMG) proteins. DNA binding studies further indicated a functional relationship of these factors to the human HMG I protein. Purification of the LAT1 factor from leaf nuclei revealed the presence of two polypeptides with molecular masses of 21 kilodaltons and 23 kilodaltons, respectively, binding the same DNA sequence with equal affinity.

Base Sequence↗
PubMed1988

Root nodule specific gene regulation: analysis of the soybean nodulin N23 gene promoter in heterologous symbiotic systems.

The nodulin N23 gene promoter was analysed in transgenic plants using the chloramphenicol acetyltransferase (CAT) coding sequence as a reporter. A 5' flanking region of less than 1 kb was sufficient for the organ-specific expression of a chimeric N23-CAT-3'lbc3 gene in root nodules formed on Lotus corniculatus and Trifolium repens after infection by their respective Rhizobium symbionts. Expression was regulated at the level of RNA in both species of transgenic plants. Promoter deletion analysis defined the 5' region required for high level expression and delimited two putative regulatory sequences involved in positive control of the N23 gene in L. corniculatus.

Cloning, Molecular↗
PubMed1987

A small family of nodule specific genes from soybean.

The primary structure of two nodule specific soybean genes are presented. The two genes code for primary products of 20.0 (nodulin 20) and 22.7 (nodulin 22) kdaltons, respectively. Both genes are related to the nodulin 23 and 44 genes. Alignment of the deduced amino acid sequences of all four genes revealed three domains of high homology interrupted by highly diverged regions due to numerous duplication and insertion events. The first conserved domain codes for a putative signal peptide, while the two others each contain four Cys residues that can be arranged in a way reminiscent of the metal binding domains present in some enzymes and in several DNA binding proteins.

Amino Acid Sequence↗
PubMed1987

Molecular cloning of lupin leghemoglobin cDNA.

Poly(A)+ RNA isolated from root nodules of yellow lupin (Lupinus luteus, var. Ventus) has been used as a template for the construction of a cDNA library. The ds cDNA was synthesized and inserted into the Hind III site of plasmid pBR 322 using synthetic Hind III linkers. Clones containing sequences specific for nodules were selected by differential colony hybridization using 32P-labeled cDNA synthesized either from nodule poly(A)+ RNA or from poly(A)+ RNA of uninfected root as probes. Among the recombinant plasmids, the cDNA gene for leghemoglobin was identified. The protein structure derived from its nucleotide sequence was consistent with known amino acid sequence of lupin Lb II. The cloned lupin Lb cDNA hybridized to poly(A)+ RNA from nodules only, which is in accordance with the general concept, that leghemoglobin is expressed exclusively in nodules.

Amino Acid Sequence↗
PubMed1986

Heme regulates the expression in Saccharomyces cerevisiae of chimaeric genes containing 5'-flanking soybean leghemoglobin sequences.

The TM1 yeast mutant was transformed with a 2 micron-derived plasmid (YEp24) which carries a chimaeric gene containing the Escherichia coli chloramphenicol acetyl transferase (CAT) gene fused to the 5'- and 3'-flanking regions of the soybean leghemoglobin (Lb) c3 gene. Expression of the chimaeric CAT gene is controlled specifically by heme at a post-transcriptional level, most likely by regulating the efficiencies of translation. Expression of another chimaeric gene consisting of the neomycin phosphotransferase (NPTII) gene fused to only the 5'-flanking region of the Lbc3 gene is regulated by heme in a similar way. Thus, in yeast, heme modulates the translation of the chimaeric mRNAs through interactions with the 5' Lbc3 non-coding region.

Acetyltransferases↗
PubMed1985

The human ubiquitin multigene family: some genes contain multiple directly repeated ubiquitin coding sequences.

Ubiquitin coding sequences were isolated from a human genomic library and two cDNA libraries. One human ubiquitin gene consists of 2055 nucleotides and codes for a polyprotein consisting of 685 amino acid residues. The polyprotein contains nine direct repeats of the ubiquitin amino acid sequence and the last ubiquitin sequence is extended with an additional valyl residue at the C-terminal end. No spacer sequences separate the ubiquitin repeats and the coding regions are not interrupted by intervening sequences. This particular gene is transcribed since cDNAs corresponding to the genomic sequence have been isolated. At least two more types of ubiquitin genes are encoded in the human genome, one coding for an ubiquitin monomer while another presumably codes for three or four direct repeats of the ubiquitin sequence. Human DNA contains many copies of the ubiquitin sequence. Ubiquitin is therefore encoded in the human genome as a multigene family.

Amino Acid Sequence↗
PubMed1984

DNA sequence and transcription of a DNA minicircle isolated from male-fertile sugar beet mitochondria.

A minicircle consisting of 1440 bp was isolated from male fertile sugar beet mitochondria. The entire nucleotide sequence of the minicircle was determined. There are several stop codons in all three reading frames and it is therefore unlikely that the minicircle codes for a polypeptide product. However, the minicircle is transcribed yielding a major RNA product consisting of 850 bases in addition to two very minor RNA products of 1100 and 950 bases, respectively. All three RNA products are transcribed from the same strand. The starting point for transcription of the major RNA product was determined by S1 nuclease protection analysis.

Base Composition↗
PubMed1984

Structure of a human gastrin gene.

A gastrin gene was isolated from a genomic library of human DNA. The human gastrin gene is about 4100 base pairs long and contains two intervening sequences. Thus, a 3500-base-pair intervening sequence is located 5 base pairs proximal to the ATG initiator codon, while a 129-base-pair intervening sequence separates the region coding for the principal hormonal form of gastrin, the heptadecapeptide, from the region coding for the major amino-terminal portion of the gastrin precursor. The 5' flanking region of the gene contains the conserved sequences, T-A-T-A-A and G-A-C-T-C-A-T-A-T, in positions similar to those of other eukaryotic genes.

Amino Acid Sequence↗
PubMed1983

Molecular cloning of human gastrin cDNA: evidence for evolution of gastrin by gene duplication.

An oligo(dT)-primed cDNA copy of the mRNA coding for the human gastrin precursor was constructed from poly(A)-containing RNA from a human pancreatic, gastrin-producing tumor (a gastrinoma). The cDNA was inserted into the Pst I endonuclease site of plasmid pBR322 by the use of the poly(dC) and poly(dG) tailing procedure. Clones containing gastrin sequences were selected by hybridization to a purified single-stranded 32P-labeled gastrin cDNA probe. This probe was constructed with gastrinoma mRNA as template. As primer for the cDNA synthesis, we used a synthetic oligonucleotide mixture, d(AG-A-A-AG-T-C-C-A-T-C-C-A), corresponding to the gastrin-specific amino acid sequence Trp-Met-Asp-Phe. In this way we determined the nucleotide sequence of the entire coding region (303 nucleotides), the entire 3' untranslated region (102 nucleotides), and 8 nucleotides of the 5' untranslated region. A striking homology between parts of the coding region suggests that evolution of the gastrin gene has involved a gene duplication.

Amino Acid Sequence↗
PubMed1982

The nucleotide sequences of two leghemoglobin genes from soybean.

We present the complete nucleotide sequences of two leghemoglobin genes isolated from soybean DNA. Both genes contain three intervening sequences in identical positions. Comparison of the coding sequences with known amino-acid sequences of soybean leghemoglobins suggest that the two genes correspond to leghemoglobin C2 and leghemoglobin C3, respectively.

Base Sequence↗