Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.30.2 (endonuclease)
 18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have developed a precise and convenient mapping technique for determining transcription start points (tsp) on cloned genomic DNA using T4 DNA polymerase. This method uses single-stranded (ss) M13 DNA and therefore is, unlike S1 and Exo VII nuclease mapping methods, independent of the restriction endonuclease sites present in the insert. Essentially the protocol involves the following steps: hybridizing an mRNA to an ss M13 vector containing an antisense genomic DNA sequence spanning the presumptive tsp (cap site); annealing a DNA primer (M13 sequencing primer) to the M13 DNA at a site on this DNA upstream from the 5' end of the mRNA on the template DNA; extending the DNA primer with T4 DNA polymerase towards the 5' end of the mRNA. Since T4 DNA polymerase will not displace the mRNA: DNA hybrid, synthesis is blocked at the first nucleotide of the mRNA molecule. The length of the extended DNA products can then be determined with single nucleotide resolution on denaturing sequencing gels in parallel with a sequencing ladder. We have used this approach to map the tsp of the mouse skeletal alpha-actin gene. The sensitivity of the method allows precise mapping of transcripts present as 0.02-0.05% of the total RNA. This method is particularly valuable for mapping the tsp of genes which are known to contain a large intron between the first and second exons. It can also be applied to map the 5' border of any given exon of a gene in an M13 vector or in other vectors that give ss DNAs.
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PMID:Mapping transcription start points on cloned genomic DNA with T4 DNA polymerase: a precise and convenient technique. 372 Dec 1

Highly purified hen oviduct progesterone receptor A subunit was tested for binding to several chicken gene DNAs. Sequence preference detected by nitrocellulose filter adsorption of [32P]DNA fragments obtained from recombinant plasmids revealed a marked retention of certain DNA fragments. About a 10-fold preference was seen for DNA fragments flanking the 5' end of the steroid-regulated genes ovalbumin and gene Y. No preference was seen with analogous DNA fragments from chicken beta-globin and alpha-actin genes. Restriction endonuclease mapping suggests the presence of multiple receptor interaction sites flanking the 5' terminus of the ovalbumin gene. One of these preferential binding sites was localized between -135 and -247 base pairs upstream from the start of transcription. This region contains an 18-base-pair A + T-rich sequence, a likely candidate for the binding site itself, because earlier studies had shown receptor A to have marked preference for A + T-rich DNA.
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PMID:DNA sequence preference of the progesterone receptor. 657 91

We have examined the relationship between chromosomal location and regulation of the two human genes encoding the sarcomeric muscle actins. The human genes encoding skeletal alpha-actin and cardiac alpha-actin are co-expressed in both human skeletal muscle and heart. We have subcloned a single-copy DNA fragment from an intervening sequence in the human cardiac alpha-actin gene and a single-copy DNA sequence from the 3' untranslated region of a human skeletal alpha-actin cDNA. Using these two gene-specific probes, we examined DNA isolated from human-mouse somatic cell hybrid lines segregating human chromosomes. We observed the segregation of restriction endonuclease-generated DNA cleavage fragments that hybridize to the two probes. The two striated muscle genes do not co-segregate and are on different autosomes. The human cardiac alpha-actin gene (ACTC) is on chromosome 15 in the q11----qter region whereas the skeletal alpha-actin gene (ACTSK) is on chromosome 1 in the p21----qter region. The co-expression of these two genes is not a function of chromosomal linkage. Neither of these muscle genes can be the primary target resulting in X-linked muscular dystrophies.
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PMID:Chromosomal location of the co-expressed human skeletal and cardiac actin genes. 658 14