Showing papers by "Applied Biosystems published in 1985"

Detection of specific sequences in nucleic acids

Abstract: The invention provides a method for diagnosis of genetic abnormalities or other genetic conditions which can be readily automated. The method is used to determine the presence or absence of a target sequence in a sample of denatured nucleic acid and entails hybridizing the sample with a probe complementary to a diagnostic portion of the target sequence (the diagnostic probe), and with a probe complementary to a nucleotide sequence contiguous with the diagnostic portion (the contiguous probe), under conditions wherein the diagnostic probe remains bound substantially only to the sample nucleic acid containing the target sequence. The diagnostic probe and contiguous probe are then covalently attached to yield a target probe which is complementary to the target sequence, and the probes which are not attached are removed. In the preferred mode, one of the probes is labeled so that the presence or absence of the target sequence can then be tested by melting the sample nucleic acid-target probe duplex, eluting the dissociated target probe, and testing for the label. In another embodiment, the testing is accomplished without first removing probes not covalently attached, by attaching a hook to the probe that is not labeled, so that the labeled target probe may be recovered by catching the hook. In both instances, the presence of both the diagnostic probe and the contiguous probe is required for the label to appear in the assay. The above method is then applied to the detection of genetic diseases.

The synthesis of oligonucleotides containing an aliphatic amino group at the 5′ terminus: synthesis of fluorescent DNA primers for use in DNA sequence analysis

Abstract: A rapid and versatile method has been developed for the synthesis of oligonucleotides which contain an aliphatic amino group at their 5' terminus. This amino group reacts specifically with a variety of electrophiles, thereby allowing other chemical species to be attached to the oligonucleotide. This chemistry has been utilized to synthesize several fluorescent derivatives of an oligonucleotide primer used in DNA sequence analysis by the dideoxy (enzymatic) method. The modified primers are highly fluorescent and retain their ability to specifically prime DNA synthesis. The use of these fluorescent primers in DNA sequence analysis will enable DNA sequence analysis to be automated.

Automated polypeptide synthesis apparatus

Abstract: An apparatus is provided for automatically constructing a polypeptide of high purity, up to 50 amino acids in length, using only single couplings. The apparatus includes an ac- ivation system for receiving protected amino acids, one kind at a time, having a common vessel (an activator vessel) in which to activate each of the amino acids. Also included is ; reaction vessel for containing a resin used in solid-phase I eptide synthesis for attaching a peptide chain thereto. A transfer system is also provided, which operates under con- rol of a computer, to transfer the activated species from the ctivation system to the reaction vessel and to transfer amino acids, reagents, gases, and solvents from one part of the apparatus to another. The activator system also includes a temperature controlled concentrator vessel in which an activator solvent is replaced by a coupling solvent to enhance the coupling of the activated species to the peptide chain in the reaction vessel. Also included in the synthesizer system is a vortexer for affecting total washing of materials in the .reaction vessel and the reaction vessel itself, an automated peptide resin sampling system, and an autodelivery system for providing individual containers of amino acid to the synthesizer in the order desired in the peptide seauence. A liquid sensor system is also included to monitor transitions between gases and liquids in specific tubes in the synthesizer in order to provide input signals to the computer system for control purposes. The computer system software which controls the operation of the synthesizer is organized according to a series of menus which allows the user of the system to select individual cycles of operation for each vessel in the synthesizer. In addition, an algorithm has been developed which provides for optimum efficiency in the production of a peptide for any given selection of cycles.

Depurination As a Yield Decreasing Mechanism in Oligodeoxynucleotide Synthesis

Abstract: The synthesis in our laboratory of very long (80–106 bases) oligodeoxynucleotides using the phosphite method of Matteucci and Caruthers in an automated DNA synthesizer indicates the true efficiency of this multiple step process. The coupling efficiency as measured by trityl cation release i s 99.5–99.8%. However the yield obtained is still below the theoretical yield calculated from the above efficiency. We have attempted t o elucidate yield decreasing mechanisms in long ol igodeoxynucleotide synthesis. Crude reaction mixtures of phosphate-deblocked oligonucleotides which had no 5′-trityl group and which were examined immediately after cleavage from the support show one major species by high resolution PAGE. Harsher hydrolysis generates a new pattern of products superimposed upon the initial ladder representing cleavage at purine residues internal to the full length oligonucleotide. In many cases the cleavage ladder is significantly more intense than the coupling ladder. Hydrolysis a t apurinic s...

A survey of interculture practices and research in Sri Lanka

Abstract: A survey on interculture research in Sri Lanka is outlined with details of species composition, spatial arrangement and justification for growing the crops as mixtures. The systems include interculture in tea, rubber and coconut plantations, spice gardens and alley cropping between leguminous tree crops. Soil conservation, generation of fuel and fodder, incentives for replanting and export diversification are some of the justifications put forward for interculture along with microclimate modification and reduction of pest incidence. The predominant expected yield criterion was some yield of associated intercrop with no reduction in tree crop yield rather than a land equivalence ratio of greater than unity. Major methodological problems were encountered due to lack of published information in mixture yields and the heterogeneity of experimental sites. Techniques for determining the biological basis of any yield advantage and the optimal spatial arrangement of intercrops were also missing.

A process for the manufacture of peptides by solid phase synthesis

Abstract: A process for the manufacture of peptides by solid phase synthesis in which successive amino acids are brought to a carboxyl-activated state and linked successively together at active sites on a solid substrate via an end carboxyl group, the amino acids being added with the amino group protected and subsequently deprotected after linkage. Successive aliquots of an alpha-amino protected form of amino acid in a non-carboxyl activated state are supplied to a common activation vessel (11) in which the amino acid is brought into a carboxyl activated state. An aliquot of an appropriate carboxyl-activated amino acid in an appropriate coupling medium is then transferred to a common reaction vessel in which the carboxyl-activated amino acid reacts with the deprotected peptide under synthesis.