Authors: Robert A. Kesterson
, Larry W. Johnson Robert A. KestersonAffiliation:
Department of Genetics, University of Alabama, Birmingham, USAFor correspondence: email@example.comBio-protocol author page: a3629
, Laura J. Lambert Larry W. JohnsonAffiliation:
Department of Genetics, University of Alabama, Birmingham, USABio-protocol author page: a3630
, Jay L. Vivian Laura J. LambertAffiliation:
Department of Genetics, University of Alabama, Birmingham, USABio-protocol author page: a3631
, Danny R. Welch Jay L. VivianAffiliation:
Department of Pathology, University of Kansas Medical Center, Kansas City, USABio-protocol author page: a3632
and Scott W. Ballinger Danny R. WelchAffiliation:
Department of Cancer Biology, University of Kansas Cancer Center, Kansas City, USAFor correspondence: firstname.lastname@example.orgBio-protocol author page: a3633
date: 10/20/2016, 18 views, 0 Q&A.
Scott W. BallingerAffiliation:
Division of Molecular and Cellular Pathology, University of Alabama, Birmingham, USAFor correspondence: email@example.comBio-protocol author page: a3634
|Brief version appeared in Cancer Res, Oct 2015 |
The mitochondrial paradigm for common disease proposes that mitochondrial DNA (mtDNA) sequence variation can contribute to disease susceptibility and progression. To test this concept, we developed the Mitochondrial-nuclear eXchange (MNX) model, in which isolated embryonic pronuclei from one strain of species are implanted into an enucleated embryo of a different strain of the same species (e.g
., C57BL/6 and C3H/HeN, Mus musculus
), generating a re-constructed zygote harboring nuclear and mitochondrial genomes from different strains. Two-cell embryos are transferred to the ostia of oviducts in CD-1 pseudopregnant mice and developed to term. Nuclear genotype and mtDNA haplotype are verified in offspring, and females selected as founders for desired MNX colonies. By utilizing MNX models, many new avenues for the in vivo
study for mitochondrial and nuclear genetics, or mito-Mendelian genetics, are now possible.
Authors: Anirban Kundu
, Sujay Paul Anirban KunduAffiliation:
Division of Plant Biology, Bose Institute, Kolkata, West Bengal, IndiaBio-protocol author page: a2047
, Amita Pal Sujay PaulAffiliation:
Division of Plant Biology, Bose Institute, Kolkata, West Bengal, IndiaBio-protocol author page: a2046
and Genotypic Technology Amita PalAffiliation:
Division of Plant Biology, Bose Institute, Kolkata, West Bengal, IndiaFor correspondence: firstname.lastname@example.orgBio-protocol author page: a2048
date: 10/20/2016, 19 views, 0 Q&A.
Genotypic Technology Private Limited, #2/13, Balaji Complex, Bangalore, IndiaBio-protocol author page: a3589
|Brief version appeared in JIPB, Jan 2014 |
This protocol describes small RNA library preparation from Vigna mungo
total RNA followed by deep sequencing and analysis for microRNA identification.
Authors: I-Chen Peng
, Alex J. Bott I-Chen PengAffiliation:
Department of Life Sciences, National Cheng Kung University, Tainan, TaiwanFor correspondence: email@example.comBio-protocol author page: a3581
and Wei-Xing Zong Alex J. BottAffiliation:
Department of Chemical Biology, Ernest Mario School of Pharmacy, Rutgers University, Piscataway, USABio-protocol author page: a3582
date: 10/5/2016, 120 views, 0 Q&A.
Wei-Xing ZongAffiliation 1:
Department of Chemical Biology, Ernest Mario School of Pharmacy, Rutgers University, Piscataway, USAAffiliation 2:
Rutgers Cancer Institute of New Jersey, New Brunswick, USAFor correspondence: firstname.lastname@example.orgBio-protocol author page: a3583
|Brief version appeared in Cell Metab, Dec 2015 |
Glutamine synthetase (GS), which catalyzes the conversion of glutamate and ammonia to glutamine, is widely distributed in animal tissues and cell culture lines. The importance of this enzyme is suggested by the fact that glutamine, the product of GS-catalyzed de novo synthesis reaction, is the most abundant free amino acid in blood (Smith and Wilmore, 1990). Glutamine is involved in many biological processes including serving as the nitrogen donor for biosynthesis, as an exchanger for the import of essential amino acids, as a means to detoxifying intracellular ammonia and glutamate, and as a bioenergetics nutrient to fuel the tricarboxylic acid (TCA) cycle (Bott et al
., 2015). The method for the assay of GS enzymatic activity relies on its γ-glutamyl transferase reaction by measuring γ-glutamylhydroxamate synthesized from glutamine and hydroxylamine, and the chromatographic separation of the reaction product from the reactants (Deuel et al
., 1978). An overview of the GS glutamyl transferase reaction can be found in Figure 1. GS activity was measured by a spectrophotometric assay at a specific wavelength of 560 nm using a microplate reader. The method is simple, and has a comparable sensitivity with those methods applying radioactively labelled substrates. This modified procedure has been applied to assay/determine GS activity in cultured cell lines including the human mammary epithelial MCF10A cells and the murine pre-B FL5.12 cells, and could be used to measure GS activity in other cell lines.
Authors: Masaki Odahara
, Takayuki Inouye Masaki OdaharaAffiliation:
Department of Life Science, College of Science, Rikkyo University, Tokyo, JapanFor correspondence: email@example.comBio-protocol author page: a3571
, Yoshiki Nishimura Takayuki InouyeAffiliation:
Department of Life Science, College of Science, Rikkyo University, Tokyo, JapanBio-protocol author page: a3572
and Yasuhiko Sekine Yoshiki NishimuraAffiliation:
Department of Botany, Graduate School of Science, Kyoto University, Kyoto, JapanBio-protocol author page: a302
date: 10/5/2016, 75 views, 0 Q&A.
Department of Life Science, College of Science, Rikkyo University, Tokyo, JapanBio-protocol author page: a3574
|Brief version appeared in Plant Journal, Nov 2015 |
In plant cells, genomic DNA exists in three organelles: the nucleus, chloroplast, and mitochondrion. Genomic DNA can be damaged by endogenous and exogenous factors, but the damaged DNA can be repaired by DNA repair systems. To quantify the extent of their repair activity of on individual genomic DNA, a PCR-based assay utilizing long amplicons is valuable for evaluable. This assay is based on the inhibitory effects of methyl methanesulfonate (MMS)-induced DNA damage on the amplicons. This assay is useful for assessing DNA double-strand repair pathways, such as homologous recombination repair, as it detects DNA double-strand breaks produced by MMS in vivo