Can I use TOP10F' competent cells for transformation of my TOPO vector that contains the ccdB gene?
Strains that contain an F plasmid, such as TOP10F', are not recommended for transformation and selection of recombinant clones in any TOPO vector containing the ccdB gene. While the F plasmid does encode the CcdA protein, which acts as an inhibitor of the CcdB gyrase-toxin protein, the half-life of the CcdA protein is shorter than that of the CcdB protein. Overexpression of the CcdB protein causes cell death when its action is not prevented by sufficient CcdA.
Have you compared in vitro transcription levels between SP6 and T7 promoters in your pCRII vectors?
No, we have not done in-house comparisons of transcription levels. It is widely known though that T7 polymerase produces more RNA than SP6 (on the order of 10-fold higher).
引用和文献 (10)
引用和文献
Abstract
Semaphorin III can function as a selective chemorepellent to pattern sensory projections in the spinal cord.
Authors:Messersmith EK, Leonardo ED, Shatz CJ, Tessier-Lavigne M, Goodman CS, Kolodkin AL
Journal:Neuron
PubMed ID:7748562
'Distinct classes of primary sensory neurons in dorsal root ganglia subserve different sensory modalities, terminate in different dorsoventral locations in the spinal cord, and display different neurotrophin response profiles. Large diameter muscle afferents that terminate in the ventral spinal cord are NT-3 responsive, whereas small diameter afferents subserving pain and ... More
A nod factor binding lectin with apyrase activity from legume roots.
Authors:Etzler ME, Kalsi G, Ewing NN, Roberts NJ, Day RB, Murphy JB
Journal:Proc Natl Acad Sci U S A
PubMed ID:10318974
'A lectin isolated from the roots of the legume, Dolichos biflorus, binds to Nod factors produced by rhizobial strains that nodulate this plant and has a deduced amino acid sequence with no significant homology to any lectin reported to date. This lectin also is an enzyme that catalyzes the hydrolysis ... More
timrit Lengthens circadian period in a temperature-dependent manner through suppression of PERIOD protein cycling and nuclear localization.
Authors:Matsumoto A, Tomioka K, Chiba Y, Tanimura T
Journal:Mol Cell Biol
PubMed ID:10330175
A fundamental feature of circadian clocks is temperature compensation of period. The free-running period of ritsu (timrit) (a novel allele of timeless [tim]) mutants is drastically lengthened in a temperature- dependent manner. PER and TIM protein levels become lower in timrit mutants as temperature becomes higher. This mutation reduces per ... More
Kainate receptor subunits expressed in single cultured hippocampal neurons: molecular and functional variants by RNA editing.
To determine the kainate receptor subunits that are found in native kainate receptors, we have applied a multiplex PCR of cDNAs reverse transcribed from mRNA harvested from single cultured hippocampal neurons after electrophysiological recording. We found that all the cells showing rapidly desensitizing currents in response to kainate express the ... More
Genetic heterogeneity of hypervariable region 1 of the hepatitis C virus (HCV) genome and sensitivity of HCV to alpha interferon therapy
Authors:Sandres K, Dubois M, Pasquier C, Payen JL, Alric L, Duffaut M, Vinel JP, Pascal JP, Puel J, Izopet J
Journal:J Virol
PubMed ID:10623727
Hepatitis C virus (HCV) populations persist in vivo as a mixture of heterogeneous viruses called quasispecies. The relationship between the genetic heterogeneity of these variants and their responses to antiviral treatment remains to be elucidated. We have studied 26 virus strains to determine the influence of hypervariable region 1 (HVR-1) ... More