Severity: Warning
Message: file_get_contents(https://...@gmail.com&api_key=61f08fa0b96a73de8c900d749fcb997acc09): Failed to open stream: HTTP request failed! HTTP/1.1 429 Too Many Requests
Filename: helpers/my_audit_helper.php
Line Number: 143
Backtrace:
File: /var/www/html/application/helpers/my_audit_helper.php
Line: 143
Function: file_get_contents
File: /var/www/html/application/helpers/my_audit_helper.php
Line: 209
Function: simplexml_load_file_from_url
File: /var/www/html/application/helpers/my_audit_helper.php
Line: 994
Function: getPubMedXML
File: /var/www/html/application/helpers/my_audit_helper.php
Line: 3134
Function: GetPubMedArticleOutput_2016
File: /var/www/html/application/controllers/Detail.php
Line: 574
Function: pubMedSearch_Global
File: /var/www/html/application/controllers/Detail.php
Line: 488
Function: pubMedGetRelatedKeyword
File: /var/www/html/index.php
Line: 316
Function: require_once
We developed phosphorylation reagents with a nitrobenzyl hydrophobic tag and used them for 5'-phosphorylation of chemically or transcriptionally synthesized RNA. The capability of hydrophobic tags to synthesize 5'-monophosphorylated RNA was evaluated based on the yield of the desired oligonucleotides, stability of protecting groups during cleavage/deprotection, separation ability in reverse-phase HPLC (RP-HPLC), and deprotection efficiency after RP-HPLC purification. The results showed that a nitrobenzyl derivative with a tert-butyl group at the benzyl position was most suitable for RNA 5'-phosphorylation. Using the developed phosphorylation reagent, we chemically synthesized 5'-phosphorylated RNA and confirmed that it could be purified by RP-HPLC and the following deprotection. In addition, we demonstrated complete chemical synthesis of minimal mRNA by chemical capping of 5'-monophosphorylated RNA. Ribonucleoside 5'-monophosphates with hydrophobic protecting groups have also been developed and used as substrates to transcriptionally synthesize 5'-phosphorylated RNA with >1000 bases. From the mixture of the by-products and the desired RNA, only 5'-monophosphorylated RNA could be effectively isolated by RP-HPLC. Furthermore, monophosphorylated RNA can be converted into circular mRNA via RNA ligase-mediated cyclization. Circular mRNA expression of nanoluciferase in cultured cells and mice. These techniques are important for the production of chemically synthesized mRNA and circular mRNA.
Download full-text PDF |
Source |
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC11551738 | PMC |
http://dx.doi.org/10.1093/nar/gkae847 | DOI Listing |
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