Description: A data set of polymorphic regions of the P. falciparum genome was mined to identify a gametocyte genotyping marker. To assess marker resolution, the number of unique haplotypes in the marker region was estimated from 205 Malawian P. falciparum whole genome sequences. Specificity of the marker for detection of mature gametocytes was evaluated using reverse transcription-polymerase chain reaction of RNA extracted from NF54 mature gametocytes and rings from a non-gametocyte-producing strain of P. falciparum. Amplicon deep sequencing was performed on experimental mixtures of mature gametocytes from two distinct parasite clones, as well as gametocyte-positive P. falciparum field isolates to evaluate the quantitative ability and determine the limit of detection of the genotyping approach.

Subject of Study

malaria parasite P. falciparum

Subject Domain

Plasmodium falciparum/genetics

Keywords

High-Throughput Nucleotide Sequencing

Malaria, Falciparum

Access via SRA
Amplicon sequence data
Accession #: PRJNA1048473
Access Restrictions: Unrestricted access
Access Instructions: Raw sequence reads are available in NCBI SRA via BioProject PRJNA1048473.
Associated Publications: Vareta J, Horstman NA, Adams M, Seydel KB, McCann RS, Cohee LM, Laufer MK, Takala-Harrison S. Genotyping Plasmodium falciparum gametocytes using amplicon deep sequencing. Malar J. 2024 Apr 6;23(1):96.
Data Type: Genomic
Equipment Used: Beckman AMPure XP bead kit
A kit for purifying and size-selecting DNA fragments using paramagnetic beads, suitable for various downstream applications.

Illumina MiSeq System
Capable of automated paired end reads.

Invitrogen MEGASchortscript T7 high yield transcription kit
A kit for efficient synthesis of RNA transcripts in vitro using T7 RNA polymerase, suitable for various applications including RNA amplification and labeling.

KAPA HiFi HotStart ReadyMix PCR kit
A PCR kit containing a hot-start DNA polymerase for robust and high-fidelity amplification of DNA targets

Luna Universal Prob One-Step RT-qPCR Kit
A kit for multiplex one-step RT-qPCR, providing sensitive and reproducible quantification of RNA targets.

Qiagen QIAamp DNA blood kit
A kit for purification of genomic DNA from blood samples, utilizing spin-column technology for efficient DNA isolation.

Qiagen QuantiTect Reverse Transcription kit
A kit designed for fast and efficient cDNA synthesis with genomic DNA elimination, ensuring high yields and sensitivity.

Qiagen RNA Protect cell reagent
A reagent for stabilizing RNA in cells and tissues, preventing degradation during storage and transport.

Qiagen RNeasy Mini kit
A kit for isolating total RNA from various sample types, utilizing silica membrane technology for efficient purification.

Qiagen RNeasy Plus 96 kit
A high-throughput kit for isolating total RNA from up to 96 samples simultaneously, offering efficient purification and high yields.

Roche Light Cycler 96 Real-time PCR machine
A real-time PCR instrument offering precise temperature control and fluorescence detection for quantitative PCR analysis.

ThermoFisher Qubit fluorometer
Benchtop fluorometer designed to accurately measure DNA, RNA, and protein quantity. Measures RNA integrity and quality.
Software Used: Primer-BLAST
An online tool provided by the NCBI for designing PCR primers, allowing users to search for primers that specifically amplify target DNA sequences while avoiding nonspecific binding.

R v4.2.3
A programming language and software environment for statistical computing and graphics.

SAS 9.4
advanced analytics, multivariate analyses, business intelligence, data management, and predictive analytics

SeekDeep
A software package utilized for processing and analyzing amplicon sequencing data, including demultiplexing, trimming, quality assessment, haplotype calling, and variant detection.
Study Type: Observational
Dataset Format(s): FASTA, FASTQ
Dataset Size: 6929 MB

Grant Support: D43TW010075/National Institutes of Health (U.S.)

U19AI129388/National Institutes of Health (U.S.)

R21AI169866/National Institutes of Health (U.S.)

UMB Data Catalog

Genotyping Plasmodium falciparum gametocytes using amplicon deep sequencing
UMB Dataset

Genotyping Plasmodium falciparum gametocytes using amplicon deep sequencing UMB Dataset

Genotyping Plasmodium falciparum gametocytes using amplicon deep sequencing
UMB Dataset