Stratagene

About: Stratagene is a based out in . It is known for research contribution in the topics: Nucleic acid & Gene. The organization has 161 authors who have published 232 publications receiving 17223 citations.

Polishing with T4 or Pfu polymerase increases the efficiency of cloning of PCR fragments

Abstract: Several of the polymerases widely used for PCR, such as Taq DNA polymerase, have been found to exhibit terminal deoxynucleotide transferase activity (1, 2, 3). This terminal transferase activity, for the most part is limited to the addition of a single nucleotide and has been named 'extendase' activity. In a widely cited study, Clark has shown that when the 3' base is a cytosine nucleotide, the Taq DNA polymerase adds predominately a single A base. This result forms the basis for the T/A cloning procedure. Although Clark's results were true for 3' C-ending fragments, Hu (3) has shown that the extended nucleotide is dependent on the specific nucleotide present at the 3' end of the synthetic double-stranded polydeoxyoligonucleotide. Different polymerases, such as Taq, T7, Klenow and Vent have different extendase characteristics with regards to which base is added at the 3' DNA ends. The polymerases derived from T4 bacteriophage and the hyperthermophilic archea Pyrococcus furiosus (available from Stratagene, La Jolla, CA) do not contain any extendase activity and in all cases were found to leave bluntended molecules.

Methods and adaptors for generating specific nucleic acid populations

Abstract: The present invention relates to methods and kits for generating or analyzing nucleic acid populations or desired nucleic acid sequences based upon replication or amplification reactions. The invention comprises methods employing adaptors ligated to nucleic acids that preferentially permit replication or amplification of desired nucleic acid sequences or preferentially eliminate undesired nucleic acids from replication or amplification. The invention also comprises adaptors useful in the methods and in kits for replicating or amplifying nucleic acids. In one embodiment, the adaptors function to protect desired nucleic acids from cleavage by a restriction enzyme while other nucleic acids are cleaved. The protected, desired nucleic acids can then be preferentially replicated or amplified. Accordingly, the invention can be used for the amplification of desired nucleic acids and the effective removal of undesired nucleic acids from a population.

Methods for detection of a nucleic acid by sequential amplification

Abstract: The invention relates to nucleic acid, flap-mediated, sequential amplification methods which permit detection of a nucleic acid target in a nucleic acid sample. The invention provides for nucleic acid synthesis dependent, flap-mediated amplification methods for sequentially producing detectable, released flaps to detect a target nucleic acid. The invention provides for both linear, and exponential nucleic acid synthesis dependent, flap-mediated, sequential amplification methods.

Spectrum of spontaneous mutations in liver tissue of lacI transgenic mice

Abstract: The advent of transgenic technology has greatly facilitated the study of mutation in animals in vivo. The Big Blue® mouse system, transgenic for the lacl gene, permits not only the quantification of mutations in different tissues but also provides for the generation of in vivo-derived mutational spectra. This report details the sequence alterations of 348 spontaneous mutations recovered from the liver of 6-8-week-old male Big Blue® mice. The spectra recovered from two strains of mice, C57B1/6 and B6C3F1, were compared and found to be very similar. The predominant mutations are G:C → A:T transitions, with 75% of these occurring at 5'-CpG-3' sequences. This mutational bias is consistent with deamination-directed mutation at methylated cytosine bases. The second most common class of mutations is G:C - T:A transversions. A significant clonal expansion of mutants was found in several animals, and this was used to make an approximate correction of the mutant frequency such that the most conservative estimate of mutation frequency is presented. The establishment of this substantial database of spontaneous mutations in the liver of Big Blue' mice is intended to serve as a reference against which mutations recovered after treatment can be compared. Environ.