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41.
DNA sequencing using rolling circle amplification and precision glass syringes in a high-throughput liquid handling system 总被引:4,自引:0,他引:4
An automated high-throughput method that employs rolling circle amplification (RCA) to generate template for large-scale DNA sequencing has been developed using liquid handling systems equipped with precision glass syringes. A protocol was designed to perform the sequencing analysis from template preparation to thermal cycle sequencing within the same vessel, thus minimizing the amount of liquid handling and transfer. The amplified DNA was directly used for cycle sequencing with no need for any purification procedures. Total RCA reaction volumes as low as 500 nL generated sufficient templates for successful sequencing. Reducing the RCA total reaction volumes by a 40-fold factor, from a total of 20 microL to 500 nL, resulted in a significant reduction in cost, from $1.25/reaction to less than $0.04/reaction. Additionally, the volume of the sequencing reactions was reduced from a total of 20 to 10 microL, thus generating a further cost advantage. This high-throughput DNA sequencing protocol maximizes the speed and precision of processing while significantly reducing the cost of amplification. 相似文献
42.
Hassan Mansouri-Torshizi Fatemeh Khosravi Arezou Ghahghaei Somaye Shahraki Sareh Zareian-Jahromi 《Journal of biomolecular structure & dynamics》2018,36(10):2713-2737
Two Zn(II) complexes of formula [Zn(bpy)(Gly)]NO3 (I) and [Zn(phen)(Gly)]NO3 (II) (where bpy = 2,2′-bipyridine, phen = 1,10-phenanthroline and Gly = glycine) were synthesized and characterized by elemental analysis, molar conductance measurements, UV–vis, FT-IR, and 1H NMR spectra. The interaction ability of these complexes with calf thymus DNA was monitored using spectroscopic methods, including UV–vis absorption spectroscopy, ethidium bromide displacement, Fourier transform infrared, and electrophoretic mobility assay. Further, the human serum albumin interactions of complexes I and II were investigated using UV–vis absorption spectroscopy, fluorescence quenching, circular dichroism, and Fourier transform infrared. The results obtained from these analyses indicated that both complexes interact effectively with CT-DNA and HSA. The binding constant (Kb), the Stern–Volmer constant (Ksv), and the number of binding sites (n) at different temperatures were determined for CT-DNA and HSA. Also, the negative ΔH° and ΔS° values showed that both hydrogen bonds and van der Waals forces played major roles in the association of CT-DNA-Zn(II) and HSA-Zn(II) complex formation. The displacement experiments suggested that Zn(II)-complexes primarily bound to Sudlow’s site II of HSA. The distance between the donor (HSA) and the acceptor (Zn(II) complexes) was estimated on the basis of the Forster resonance energy transfer (FRET) and the alteration of HSA secondary structure induced by the compounds were confirmed by FT-IR spectroscopy. The complexes follow the binding affinity order of I > II with DNA and II > I with HSA. Finally, Antibacterial activity of complexes I and II have been screened against gram positive and gram negative bacteria. 相似文献
43.
Karamzade Arezou Babaei Meisam Saberi Mohammad Golchin Neda Khalil Nejad Sani Banaei Aysun Eshaghkhani Yeganeh Golchehre Zahra Keramatipour Mohammad 《Molecular biology reports》2021,48(6):5339-5345
Molecular Biology Reports - Joubert syndrome (JS) is a rare inherited neurodevelopmental condition characterized by hypotonia, ataxia, developmental delay, abnormal eye movements, neonatal... 相似文献
44.
Amyloids are insoluble fibers which arise from inappropriately folded versions of proteins and have been associated with the pathology of many neurodegenerative diseases. α-Casein is one of the major components of the casein family which is known to show chaperone-like activity. Glycerol is a polyol compound which acts as a chemical chaperone to increase protein stability and inhibit protein aggregation. In this study, the effect of arginine and glycine on the chaperone ability of α-casein and glycerol against order aggregation of κ-casein was investigated and compared. We found that these additives reduced the chaperone ability of α-casein against the amyloid formation of κ-casein, especially in the presence of arginine. Importantly, our results show that the chaperone action of glycerol is enhanced in the presence of both arginine and glycine. Accordingly, our results suggest that these small molecules associated with glycerol, especially glycine, should be considered as a mechanism for the treatment of amyloid disease. 相似文献
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Amyloid fibrils arise from the slow aggregation of intermediately folded protein states. In this study the kinetics of the
protein fibril formation of α-lactalbumin and its prevention by αS-casein in the presence and absence of the crowding agent,
dextran (68 kDa), have been compared using a thioflavin T binding assay. It was found that αS-casein, a molecular chaperone
found in bovine milk, is a potent in vitro inhibitor of α-lactalbumin fibrillization. The effect of αS-casein in preventing
fibril formation was significant, although less than it is in the absence of the crowding agent, dextran. The interaction
between the chaperone and the α-lactalbumin and structural change in the target protein are also shown using intrinsic fluorescence
intensity, an ANS binding assay, CD spectroscopy and size-exclusion HPLC. In summary, α-casein interacts with α-lactalbumin
and prevents amyloid formation but not as well as it does when the crowding agent, dextran, not present. 相似文献