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In JoVE (1)
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Articles by Harini Ravi in JoVE
JoVE General
ה-DNA מבוססי פרוטוקול זיהוי דגים המינים
פרסום זה מתאר כיצד להשתמש מיני דגים Agilent מערכת זיהוי כדי לזהות את המין של דג על ידי הפקת DNA וביצוע PCR וניתוח RFLP.
Other articles by Harini Ravi on PubMed
Aip1 and Cofilin Promote Rapid Turnover of Yeast Actin Patches and Cables: a Coordinated Mechanism for Severing and Capping Filaments
Rapid turnover of actin structures is required for dynamic remodeling of the cytoskeleton and cell morphogenesis, but the mechanisms driving actin disassembly are poorly defined. Cofilin plays a central role in promoting actin turnover by severing/depolymerizing filaments. Here, we analyze the in vivo function of a ubiquitous actin-interacting protein, Aip1, suggested to work with cofilin. We provide the first demonstration that Aip1 promotes actin turnover in living cells. Further, we reveal an unanticipated role for Aip1 and cofilin in promoting rapid turnover of yeast actin cables, dynamic structures that are decorated and stabilized by tropomyosin. Through systematic mutagenesis of Aip1 surfaces, we identify two well-separated F-actin-binding sites, one of which contributes to actin filament binding and disassembly specifically in the presence of cofilin. We also observe a close correlation between mutations disrupting capping of severed filaments in vitro and reducing rates of actin turnover in vivo. We propose a model for balanced regulation of actin cable turnover, in which Aip1 and cofilin function together to "prune" tropomyosin-decorated cables along their lengths. Consistent with this model, deletion of AIP1 rescues the temperature-sensitive growth and loss of actin cable defects of tpm1Delta mutants.
Rapid Quantification of DNA Libraries for Next-generation Sequencing
The next-generation DNA sequencing workflows require an accurate quantification of the DNA molecules to be sequenced which assures optimal performance of the instrument. Here, we demonstrate the use of qPCR for quantification of DNA libraries used in next-generation sequencing. In addition, we find that qPCR quantification may allow improvements to current NGS workflows, including reducing the amount of library DNA required, increasing the accuracy in quantifying amplifiable DNA, and avoiding amplification bias by reducing or eliminating the need to amplify DNA before sequencing.
