Globin Reduction PNA

Globin Reduction PNA

Globin Reduction PNA is an efficient and reliable method provided by Creative Peptides for eliminating globin mRNA interference in RNA sequencing applications.

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Introduction

Introduction to PNA

PNA (peptide nucleic acid) is a synthetic chemical similar to DNA and RNA. The backbone of PNA is composed of repeating N-2-(aminoethyl)-glycine units that are combined by peptide bonds. The base and backbone are connected by a methylene carboxyl bond. PNA has the following characteristics.

• PNA has no phosphate groups such as those found on nucleic acids, so there is a lack of electrical repulsion between PNA and DNA and RNA, so the binding strength between PNA and DNA is greater.
• PNA is neither a polypeptide nor a nucleic acid. Therefore, PNA is not readily hydrolyzed by proteases or nucleases, and PNA is quite stable in vivo and in vitro.
• There is a strong pairing specificity and high thermal stability between PNA and nucleic acid bases.

Globulin-reduced PNA is an important application area.

Globin mRNA

Globulin mRNA accounts for more than 70% of total mRNA in blood, which will severely affect the amplification of non-globulin mRNA. Transcriptome analysis of whole blood RNA by sequencing holds promise for identification and tracking biomarkers. However, the high globin mRNA (gmRNA) content of erythrocytes hinders whole blood and water-coat analysis. With the removal of globin mRNA, RNA libraries are more amenable to biological experiments such as gene cloning and transcriptome sequencing.

Globin Reduction PNA

The primary function of Globin Reduction PNA is to reduce the effect of Globin mRNA in blood on whole blood transcriptome sequencing. During reverse transcription and subsequent PCR amplification, PNA acts as a sequence-specific blocker, preventing the amplification of globin mRNA, thereby making it easier for non-globin mRNA to be amplified.

Reduction of globin PNA improves detection sensitivity by reducing the amount of cDNA produced from globin mRNA in the first-strand cDNA synthesis step. To achieve this, PNA oligonucleotides are added to the cellular RNA samples. These PNAs are complementary to the human globin mRNA transcript and anneal to the globin mRNA transcript in the primer hybridization step. In the subsequent first-strand cDNA synthesis reaction, the PNAs bound to the globin mRNA block reverse the transcription of the corresponding transcripts. Consequently, the amount of cDNA produced from globin mRNA and subsequently synthesized cRNA is reduced, allowing for sensitive and unbiased gene expression analysis of other cells found in the blood.

Sequence information of common Globin-reduced PNAs

• Globin- Reduction PNA 1: Ac- TAA CGG TAT TTG GAG
• Globin-Reduction PNA 2: Ac- GTA GTT GGA CTT AGG
• Globin-Reduction PNA 3: Ac- GCC CTT CAT AAT ATC
• Globin- Reduction PNA 4: Ac- ATC CAG ATG CTC AAG

The sequences above are written from the N-terminal (5' end) to the C-terminal (3' end). The 5' end is acetylated (Ac).

References

• Egholm M; et al. PNA hybridizes to complementary oligonucleotides obeying the Watson-Crick hydrogen-bonding rules. Nature. 1993, 365(6446): 566-568.
• Dwaine, A; et al Novel antisense and peptide nucleic acid strategies for controlling gene expression. Biochemistry. 2002, 41(14): 4503–4510.
• Nielsen P E. Peptide nucleic acid. A molecule with two identities. Accounts of Chemical Research. 1999, 32(7).
• Lori A; et al. Functional identity of genes detectable in expression profiling assays following globin mRNA reduction of peripheral blood samples. Clinical Biochemistry. 2007.
• Krjutkov K; et al. Globin mRNA reduction for whole-blood transcriptome sequencing. Scientific Reports. 2016, 6: 31584.

For Research Use Only. Not For Clinical Use.