Andrea Jeffrey

Bio: Andrea Jeffrey is an academic researcher from Harvard University. The author has contributed to research in topics: Base pair & Repressor. The author has an hindex of 10, co-authored 12 publications receiving 3929 citations.

Nucleotide sequence of the rightward operator of phage lambda

Abstract: The sequence of 72 base pairs of the rightward operator (O-R) of bacteriophage lambda is presented as determined with simple and rapid methods for direct DNA sequencing The sequence of an operator mutant is also described The methods are of general use in sequencing DNA fragments with unique 5' ends up to 50 base pairs in length Previous experiments have shown that this operator contains multiple sites recognized by the lambda phage repressor We believe we have identified three of these sites

Chain length determination of small double- and single-stranded DNA molecules by polyacrylamide gel electrophoresis

Abstract: We describe the use of polyacrylamide gel electrophoresis to estimate chain lengths of double- and single-stranded DNA molecules in the size range 20-1000 base pairs (or nucleotides). Double-stranded DNA molecules of known length produced either by organic synthesis or by restriction endonuclease digestion of viral DNAs were used as standards. The relative electrophoretic mobilities of these standards were examined on both nondenaturing (aqueous) polyacrylamide gels and on denaturing gels containing 7 M urea or 98% formamide. Electrophoretic mobility of DNA is a linear function of the log of molecular weight if appropriate conditions are used, although exceptions are noted. Chain lengths can be conveniently estimated by using as standards bacteriophage gamma DNA restriction fragments or commercially available tracking dyes.

Sequencing end-labeled DNA with base-specific chemical cleavages.

Abstract: Publisher Summary This chapter discusses the sequencing end-labeled DNA with base-specific chemical cleavages. In the chemical DNA sequencing method, one end-labels the DNA, partially cleaves it at each of the four bases in four reactions, orders the products by size on a slab gel, and then reads the sequence from an autoradiogram by noting which base-specific agent cleaved at each successive nucleotide along the strand. This technique sequences the DNA made in and purified from cells. No enzymatic copying in vitro is required, and either single- or double-stranded DNA can be sequenced. Most chemical schemes that cleave at one or two of the four bases involve three consecutive steps: modification of a base, removal of the modified base from its sugar, and DNA strand scission at that sugar. Base-specific chemical cleavage is only one step in sequencing DNA. The chapter presents techniques for producing discrete DNA fragments, end-labeling DNA, segregating end-labeled fragments, extracting DNA from gels, and the protocols for partially cleaving it at specific bases using the chemical reactions. The chapter also discusses the electrophoresis of the chemical cleavage products on long-distance sequencing gels and a guide for troubleshooting problems in sequencing patterns.

From molecular to modular cell biology.

Abstract: Cellular functions, such as signal transmission, are carried out by 'modules' made up of many species of interacting molecules Understanding how modules work has depended on combining phenomenological analysis with molecular studies General principles that govern the structure and behaviour of modules may be discovered with help from synthetic sciences such as engineering and computer science, from stronger interactions between experiment and theory in cell biology, and from an appreciation of evolutionary constraints

Purification of mouse immunoglobulin heavy-chain messenger RNAs from total myeloma tumor RNA.

Abstract: A procedure is described for the large-scale purification of light (L) and heavy (H) chain mRNAs from plasmacytomas produced in mice. Intact RNA is selectively precipitated in high yield from frozen tumors homogenized in 3 M LiCl and 6 M urea. L and H-chain mRNAs were purified by oligo(dT)-cellulose chromatography and either sucrose gradient centrifugation in conditions preventing aggregation or by means of high-resolution preparative gel electrophoresis under non-denaturing conditions. γ2a and α H-chain mRNAs sedimented as major components at 15.5 S and 16.5 S respectively, when L-chain mRNAs sedimented as 12-S species. H-chain mRNAs isolated by continuous elution during preparative gel electrophoresis were completely separated from both L-chain mRNA and residual 18-S rRNA, and migrated as single components of 1900 ± 50 nucleotides on analytical denaturing gels. The partially purified H-chain mRNAs were translated into major components of molecular weights of 56000 (γ2a) and 60000 (α) in an mRNA-dependent rabbit reticulocyte lysate, whereas L-chain mRNAs yielded polypeptides of molecular weights of 25000 (λ) and 27000 (к). Up to 95% of the translation products directed by the purified mRNAs were immunoprecipitated using specific antisera. The purity of L and H-chain mRNAs was assessed by hybridization of corresponding cDNAs with excess recombinant plasmid DNA. The results indicated a minimum purity of 47% (γ2a), 62% (α), for H-chain mRNAs and 60% (к), for L-chain mRNAs.

Transformation of yeast

Abstract: A stable leu2- yeast strain has been transformed to LEU2+ by using a chimeric ColE1 plasmid carrying the yeast leu2 gene. We have used recently developed hybridization and restriction endonuclease mapping techniques to demonstrate directly the presence of the transforming DNA in the yeast genome and also to determine the arrangement of the sequences that were introduced. These studies show that ColE1 DNA together with the yeast sequences can integrate into the yeast chromosomes. This integration may be additive or substitutive. The bacterial plasmid sequences, once integrated, behave as a simple Mendelian element. In addition, we have determined the genetic linkage relationships for each newly introduced LEU2+ allele with the original leu2- allele. These studies show that the transforming squences integrate not only in the leu2 region but also in several other chromosomal locations.