bioRxiv Science⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.1101/2024.08.04.606550

Optimized semi-specific PCR amplification using arbitrarily degenerate primer for genome-wide genotyping and its application in peanut genetic diversity study

Zhao, S.·Wang, Y.·Zhang, X.·Xie, S.·Chen, H.·Yan, Y.·Gu, J.·Chen, P.·Chang, Y.·Xu, Z.
View original source ↗Export metadata ↓

Abstract

Cultivated peanut (Arachis hypogaea L.) is an important legume crop with a large genome size but a limited genetic diversity. Evaluating the genetic variation among diverse peanut germplasms using genome-wide molecular markers is an effective strategy to explore its genetic diversity and thereby facilitate peanut improvement. In this study, we introduced a novel whole-genome genotyping (WGG) technique named dRAPD-seq (degenerate Random Amplification Polymorphic DNA and sequencing), which relies on semi-specific PCR amplification by arbitrarily degenerate (AD) primer and next-generation sequencing (NGS), and demonstrated its robust reproducibility and high accuracy. Subsequently, we applied dRAPD-seq to investigate the genetic relatedness within a population of 101 diverse peanut accessions and identified a low genetic diversity among these accessions. Our phylogenetic tree, population structure analyses, and principal component analysis (PCA) indicated that this population could be clustered into three subpopulations, largely corresponding to three botanical types. In conclusion, this research not only introduced a cost-effective and easy-to-conduct WGG method but also provided valuable insights for utilizing these peanut accessions in future breeding programs.

Explore related subjects

genomicsPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Zhao, S., Wang, Y., Zhang, X., Xie, S., Chen, H., Yan, Y., Gu, J., Chen, P., Chang, Y., Xu, Z.. 2024-08-07. Optimized semi-specific PCR amplification using arbitrarily degenerate primer for genome-wide genotyping and its application in peanut genetic diversity study. https://doi.org/10.1101/2024.08.04.606550

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

Structural variation in repeat elements is widespread in normal human tissues and in tumorigenesis

Somatic mosaicism contributes to genomic variation, yet postzygotic structural variants remain under-characterized. We performed long- and short-read WGS from multiple individuals (n=47 normal tissues; n=168 samples) and identified mosaic structural variants in all individuals and germ layers, impacting a median 285.2 kb/genome. Nearly half of breakpoints were independently validated, with tissue distributions reflecting both early and late developmental origins. Most mosaic variants were repeat-mediated and 8.3% overlapped functional elements, an enrichment compared to germline variants. To extend these analyses in samples where long-read sequencing is infeasible, we measured repeat alterations from short-read sequencing, recapitulating mosaic tissue-specific differences. We characterized tumor- and tissue- specific variation in repeats across 15 cancer types and found tumor-related repeat variation to be similar in scale to that of normal mosaic variation. Tracking repeat changes in cell-free DNA provided a noninvasive approach for tumor monitoring. Our analyses revealed widespread repeat-driven structural variation in health and disease.

genomics↗
bioRxiv2026

RNA isoform-resolved multiplexed sequencing with bioorthogonal barcoding

RNA isoform dysregulation drives disease pathogenesis and is the target of FDA-approved splice-switching therapeutics. However, multiplexed sequencing methods discard splice junction information because only 3' termini are barcoded and counted. Here, we repurpose acylation and click chemistries to conjugate bioorthogonal barcodes (bobcodes) directly onto multiple internal positions along cellular RNAs. Bobcoded RNAs from multiple samples are pooled for multiplexed cDNA synthesis, during which reverse transcriptase switches from each RNA template onto its tethered bobcode with greater than 99% accuracy in species mixing experiments. Bobcode attachment intervals set cDNA insert sizes without a library fragmentation step, and priming with poly(dT) or random hexamers selects between 3'-end counting and full-length isoform capture. A bioorthogonal barcode-sequencing (BOB-seq v0.1) drug screen identifies transcriptome-wide on- and off-target RNA splicing effects and outperforms existing multiplexing RNA sequencing methods in workflow simplicity, sample-to-sample variability, and barcoding accuracy. Bobcodes add isoform resolution to scalable multiplexed RNA sequencing.

genomics↗
bioRxiv2026

Structural polymorphism and population-variable coding capacity of HERV-K(HML-2) in human pangenomes

Approximately 8% of the human genome is derived from ancient retroviral infections. The most recently integrated of these endogenous retroviruses is the HERV-K(HML-2) clade, whose expression has been associated with cancer, amyotrophic lateral sclerosis, and embryogenesis. Studies of HERV expression, particularly HML-2, have relied predominantly on short-read sequencing. However, the high similarity among HML-2 proviruses prevents many short reads from being assigned uniquely to individual loci. We therefore compared haplotype-resolved long-read genome assemblies from 292 donors to resolve variation in proviral structure and coding capacity. Several loci previously thought to be fixed were structurally polymorphic. Tandem arrays occurred at 13 loci and contained up to six proviral copies in a single array. At 8q11.23, we identified a previously undescribed full-length provirus in one haplotype. All 583 other haplotypes carried a solo-LTR. We found that standard reference genomes failed to represent the coding capacity retained in many individuals, whose proviruses contained intact open reading frames despite disruptive mutations in the reference sequences. Short-read genotypes left 32.5% of the tested donor-variant pairs unresolved at sites associated with viral reading frames. These findings show why HML-2 expression must be interpreted in the context of the structural and coding alleles each individual carries.

genomics↗