Efficient Cloning and Sequence Validation of Repetitive and High GC-Content Short Hairpin RNAs

Chilamkurthy, Ramadevi; White, Adam A; Pater, Adrian A; Jensik, Philip J.; Gagnon, Keith T.

doi:10.1089/hum.2021.273

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2024

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Cited by 2 publications

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“…To generate small RNA expression vectors, we cloned small RNA duplexes ( Table S1 ) into a U6 promoter-driven lentiviral vector expressing a separate EGFP-puromycin reporter 24 , 25 ( Figure S1 A). Small RNAs were expressed from two core constructs, an shRNA 24 ( Figure 1 B) or a miRNA ( Figure 1 C) scaffold.…”

Section: Resultsmentioning

confidence: 99%

Sequencing-guided design of genetically encoded small RNAs targeting CAG repeats for selective inhibition of mutant huntingtin

Parasrampuria,

White,

Chilamkurthy

et al. 2024

Molecular Therapy - Nucleic Acids

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Section: Resultsmentioning

confidence: 99%

Sequencing-guided design of genetically encoded small RNAs targeting CAG repeats for selective inhibition of mutant huntingtin

Parasrampuria,

White,

Chilamkurthy

et al. 2024

Molecular Therapy - Nucleic Acids

Self Cite

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Visual codon: a user-friendly Python program for viewing and optimizing gene GC content

Lin,

Xu,

Huang

et al. 2024

PeerJ

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Due to the codon bias of different species, codon optimization is usually carried out in the process of heterologous protein expression. At present, there are a variety of codon optimization tools. However, the optimized sequences may still have high or low points of local guanine and cytosine (GC) content, which is not conducive to the primer design of gene subcloning, and also makes it difficult to perform the experiment of synthesizing the whole gene with DNA fragments by polymerase chain reaction (PCR) reaction. In this study, we present a stand-alone software written in Python, with which users can manually check and adjust the GC content of sequence-optimized genes. The software takes the codon frequency of Escherichia coli as default and can work with other species as well. It provides a Graphical User Interface (GUI) interface, which allows users to change codons and intuitively see the effect of codon changes on local GC content. Our program brings convenience for the optimization of gene GC content and the subsequent gene cloning experiments.

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Efficient Cloning and Sequence Validation of Repetitive and High GC-Content Short Hairpin RNAs

Cited by 2 publications

References 42 publications

Sequencing-guided design of genetically encoded small RNAs targeting CAG repeats for selective inhibition of mutant huntingtin

Sequencing-guided design of genetically encoded small RNAs targeting CAG repeats for selective inhibition of mutant huntingtin

Visual codon: a user-friendly Python program for viewing and optimizing gene GC content

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