| 1. |
- Klionsky, Daniel J., et al.
(författare)
-
Guidelines for the use and interpretation of assays for monitoring autophagy in higher eukaryotes
- 2008
-
Ingår i: Autophagy. - : Landes Bioscience. - 1554-8627 .- 1554-8635. ; 4:2, s. 151-175
-
Forskningsöversikt (refereegranskat)abstract
- Research in autophagy continues to accelerate,1 and as a result many new scientists are entering the field. Accordingly, it is important to establish a standard set of criteria for monitoring macroautophagy in different organisms. Recent reviews have described the range of assays that have been used for this purpose.2,3 There are many useful and convenient methods that can be used to monitor macroautophagy in yeast, but relatively few in other model systems, and there is much confusion regarding acceptable methods to measure macroautophagy in higher eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers of autophagosomes versus those that measure flux through the autophagy pathway; thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from fully functional autophagy that includes delivery to, and degradation within, lysosomes (in most higher eukaryotes) or the vacuole (in plants and fungi). Here, we present a set of guidelines for the selection and interpretation of the methods that can be used by investigators who are attempting to examine macroautophagy and related processes, as well as by reviewers who need to provide realistic and reasonable critiques of papers that investigate these processes. This set of guidelines is not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to verify an autophagic response.
|
|
| 2. |
- Creusot, F., et al.
(författare)
-
The CIC YAC library: Sizing of the clones and determination of clones carrying repeated DNA sequences.
- 1995
-
Ingår i: The Plant Journal. - 0960-7412 .- 1365-313X. ; 8:5, s. 763-770
-
Tidskriftsartikel (refereegranskat)abstract
- A new Arabidopsis thaliana (ecotype Columbia) genomic library has been constructed in Yeast Artificial Chromosomes: the CIC library (for CEPH, INRA and CNRS). Optimization of plant culture conditions and protoplast preparation allowed the recovery of large amounts of viable protoplasts. Mechanical shearing of DNA was minimized by isolation of DNA from protoplasts embedded in agarose. Cloning of large inserts was favored by including two successive size fractionation steps (after partial EcoRI digestion and after ligation with the vector arms), which selected DNA fragments larger than 350 kb. The library consists of 1152 clones with an average insert size of 420 kb. Clones carrying chloroplast DNA and various nuclear repeated sequences have been identified. Twenty-one per cent of the clones are found to contain chloroplast DNA. Therefore, the library represents around four nuclear genome equivalents. The clones containing 5S rDNA genes, 18S-25S rDNA sequences and the 180 bp paracentromeric repeated element account for 3.6%, 8.9% and 5.8%, respectively. Only one clone was found to carry the 160 bp paracentromeric repeated element. Given the smaller size of clones carrying Arabidopsis repeated DNA, the average size of remaining clones is around 480 kb. The library was screened by PCR amplification using pairs of primers corresponding to sequences dispersed in the genome. Seventy out of 76 pairs of primers identified from one to seven YAC clones. Thus at least 92% of the genome is represented in the CIC library. The survey of the library for clones containing unlinked DNA sequences indicates that the proportion of chimeric clones is lower than 10%.
|
|
| 3. |
- Bukartyk, Marta М., et al.
(författare)
-
Preparation and research of properties of combined alginate/gelatin hydrogels
- 2022
-
Ingår i: Journal of Chemistry and Technologies. - Ukraine : Oles Honchar Dnipropetrovsk National University. - 2663-2934 .- 2663-2942. ; 30:1, s. 11-20
-
Tidskriftsartikel (refereegranskat)abstract
- The work presents the studies on the synthesis of hydrogel material based on the natural biopolymers (sodium alginate and gelatin) for medical applications. Sodium alginate and gelatin are biocompatible, non-toxic, biodegradable polymers and renewable raw materials. Combined alginate gelatin hydrogels are formed due to the formation of a hydrogel network by simultaneous cross-linking of calcium ions of sodium alginate macromolecules, gelatin, and macro-chains of rarely cross-linked polyacrylic acid. The optimal synthesis conditions (reagent concentrations, the molar ratio of calcium ions to the number of carboxyl groups Ca2+/COOH-) of the combined hydrogels with satisfactory physicochemical and mechanical properties are determined. The dependences of the mechanical properties of alginate-gelatin hydrogels on the degree of swelling indicate a wide range of their satisfactory performance characteristics. The studies on sorption and release of analgesics (lidocaine and novocaine) show long-term release of drugs and allow predicting the possibility of their prolonged delivery. In vitro cytotoxicity analysis proved the absence of toxic effect on living cells. The results suggest that the obtained combined alginate-gelatin hydrogels are a promising material for producing hydrogel dressings for wound care.
|
|
