The isolation of translatable poly(A)+mRNA from the slime glands of the Pacific hagfish, Eptatretus stouti, is not possible by the commonly used procedures because of the viscous slime that is formed when the contents of the glands are hydrated. This paper reports on a procedure developed to overcome this problem. Briefly, the tissue was powdered in liquid nitrogen, mixed with sodium lauroylsarcosine and proteinase K and lyophilized. The lyophilized powder was then mixed with 0.3 mm diameter glass beads, thoroughly ground and wetted with buffer and digested at 37 degrees C. The RNA from the digest was recovered by ultracentrifugation through a CsCl cushion. Further purification of the RNA was accomplished by the usual methods with slight modifications.
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http://dx.doi.org/10.1080/00327488808062513 | DOI Listing |
J Pharm Biomed Anal
January 2025
Analytical Research and Development, Merck & Co., Inc., Rahway, NJ 07065, United States. Electronic address:
The dynamic landscape of mRNA technology highlights the need for innovative quality control (QC) strategies. In this study, we described an efficient one-step digestion approach for concurrent generation of 5'- and 3'-end fragments, enabling simultaneous mRNA capping and poly(A) tail analysis. Tailored 10-23-type DNAzymes, designed from 5'- and 3'-Untranslated Regions (UTRs), selectively cleaved mRNA to release both the 5'-Capped or uncapped short fragments and 3'-Poly(A) tail cleavage products.
View Article and Find Full Text PDFPharmaceutics
December 2024
Gennova Biopharmaceuticals Ltd., ITBT Park, Hinjawadi Phase 2 Rd, Hinjewadi Rajiv Gandhi Infotech Park, Hinjawadi, Pune 411057, India.
: The nucleic acid-based product (NAP) portfolio is expanding continuously and provides safer curative options for many disease indications. Nucleic acid-based products offer several advantages compared to proteins and virus-based products. They represent an emerging field; thus, their quality control and regulatory landscape is evolving to ensure adequate quality and safety.
View Article and Find Full Text PDFBiology (Basel)
January 2025
Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences (ICBFM SB RAS), 8, Lavrentiev Avenue, Novosibirsk 630090, Russia.
Poly(A) polymerase (PAP 1) from is the primary enzyme responsible for synthesizing poly(A) tails on RNA molecules, signaling RNA degradation in bacterial cells. In vitro, PAP 1 is used to prepare libraries for RNAseq and to produce mRNA vaccines. However, PAP 1's toxicity and instability in low-salt buffers complicate its expression and purification.
View Article and Find Full Text PDFRespir Res
January 2025
Department of Obstetrics and Gynecology, C.S. Mott Center for Human Growth and Development, School of Medicine, Wayne State University, 275 E Hancock St, Rm 195, Detroit, MI, 48201, USA.
Current fetal alcohol spectrum disorders (FASD) studies primarily focus on alcohol's actions on the fetal brain although respiratory infections are a leading cause of morbidity/mortality in newborns. The limited studies examining the pulmonary adaptations in FASD demonstrate decreased surfactant protein A and alveolar macrophage phagocytosis, impaired differentiation, and increased risk of Group B streptococcal pneumonia with no study examining sexual dimorphism in adaptations. We hypothesized that developmental alcohol exposure in pregnancy will lead to sexually dimorphic fetal lung morphological and immune adaptations.
View Article and Find Full Text PDFChembiochem
January 2025
Modality Research Laboratories 1, Research Unit, Research Division, Kyowa Kirin Co., Ltd., 3-6-6 Asahi, Machida, Tokyo, 194-8533, Japan.
Position-specific nucleoside sugar modifications have been shown to improve the translational activity and stability of chemically synthesized mRNA. For pharmaceutical applications of chemically modified mRNAs, a rapid purification methodology is imperative to identify the optimal modification pattern. However, while the chemical synthesis of mRNAs can be accomplished by splint ligation of oligonucleotide fragments, the current purification method for ligated mRNAs based on denaturing polyacrylamide gel electrophoresis tends to be time consuming.
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