Barcode System Research Articles

Abstract 5544: Identification, isolation and molecular characterization of drug-resistant sub-populations of pancreatic cancer cells

Abstract Pancreatic ductal adenocarcinoma (PDAC) has a poor prognosis, since most PDAC tumors develop resistance to the standard of care (SoC) treatments like chemotherapy, radiation, and targeted therapies. All tumors consist of multiple, genetically related subpopulations of cancer cells that evolve in parallel and display heterogeneity at genomic, epigenetic, or phenotypic levels. As cancer develops, some subpopulations of cancer cells may show faster growth, increased metastatic potential, and resistance toward SoC treatment. A key challenge in PDAC management is understanding chemoresistance driven by cancer subpopulation dynamics. This would broaden our understanding of tumor adaptation to treatments and guide targeting resistant subpopulations to improve therapeutic outcomes. In this study, we identified PDAC cell subpopulations resistant to chemotherapy using our DNA barcoding system B-GLI (barcode-guide lineage isolation). The B-GLI system leverages a highly complex DNA barcode library and CRISPR activation (CRISPRa) to trace and isolate sub-lineages within heterogeneous cell populations by their DNA barcodes. PDAC cell lines PANC-1 and Mia-PaCa-2 were barcoded with the B-GLI barcode library, which consist of two optimized guide RNA binding sites in each barcode. After DNA barcoding, we performed treatments with two SoC chemotherapeutics, gemcitabine + paclitaxel and FOLFIRINOX, to induce a selection pressure in the PDAC cell lines, and then identified the resistant subpopulations through differentially represented barcodes and sequencing. Furthermore, we designed barcode-specific single guide RNAs (sgRNAs) targeting the resistant subpopulations and activated the expression of the puromycin resistance gene by CRISPRa. This allowed us to enrich and isolate chemoresistant subpopulations by puromycin treatment. After having isolated the resistant PDAC subpopulations, we performed ATAC-seq and RNA-seq for molecular characterization of the resistant and parental cell populations, along with phenotypic drug screening with 384 compounds to identify novel treatment vulnerabilities. The compound testing confirmed that the resistant cells were less sensitive to the two SoC treatments, as well as identified selective sensitivity to specific compounds. ATAC-seq and RNA-seq data will allow us to decode whether and how chromatin structure and transcriptomic changes are associated with the treatment resistance, and the confirmation screen of the drug sensitivities will identify potential compounds for future pre-clinical and clinical testing in PDAC models and patients with resistant disease. In summary, this study will identify targeted treatment alternatives, accompanied by molecular biomarkers for chemotherapy resistant PDAC. Citation Format: Subhendu Roy Choudhury, Shixiong Wang, Yevhen Akimov, Katarina Willoch, Biswajyoti Sahu, Alfonso Urbanucci, Thomas Fleischer, Tero Aittokallio. Identification, isolation and molecular characterization of drug-resistant sub-populations of pancreatic cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5544.

Precise Lineage Tracking Using Molecular Barcodes Demonstrates Fitness Trade-offs for Ivermectin Resistance in Nematodes.

A fundamental tenet of evolutionary genetics is that the direction and strength of selection on individual loci varies with the environment. Barcoded evolutionary lineage tracking is a powerful approach for high-throughput measurement of selection within experimental evolution that to date has largely been restricted to studies within microbial systems, largely because the random integration of barcodes within animals is limited by physical and molecular protection of the germline. Here, we use the recently developed TARDIS barcoding system in Caenorhabditis elegans (Stevenson et al., 2023) to implement the first randomly inserted genomic-barcode fitness experiment within an animal model and use this system to precisely measure the influence of the concentration of the anthelmintic compound ivermectin on the strength of selection on an ivermectin resistance cassette. The combination of the trio of knockouts in neuronally expressed GluCl channels, avr-14, avr-15, and glc-1, has been previously demonstrated to provide resistance to ivermectin at high concentrations. Varying the concentration of ivermectin in liquid culture allows the strength of selection on these genes to be precisely controlled within populations of millions of individuals, with the frequency of each barcode then being measured at multiple time points via sequencing at deep coverage and used to estimate the fitness of the individual lineages in the population. The mutations display a high cost to resistance at low concentrations, rapidly losing out to wildtype genotypes, but the balance tips in their favor when the ivermectin concentration exceeds 2nM. This trade-off in resistance is likely generated by a hindered rate of development in resistant individuals. Our results demonstrate that C. elegans can be used to generate high precision estimates of fitness using a high-throughput barcoding approach to yield novel insights into evolutionarily and economically important traits.

Design and synthesis of a new highly efficient adjustable Ln-MOF for fluorescence sensing and information encryption.

Design and synthesis of a new highly efficient adjustable Ln-MOF for fluorescence sensing and information encryption.

Deep-tissue transcriptomics and subcellular imaging at high spatial resolution.

Limited color channels in fluorescence microscopy have long constrained spatial analysis in biological specimens. We introduce cycle hybridization chain reaction (cycleHCR), a method that integrates multicycle DNA barcoding with HCR to overcome this limitation. cycleHCR enables highly multiplexed imaging of RNA and proteins using a unified barcode system. Whole-embryo transcriptomics imaging achieved precise three-dimensional gene expression and cell fate mapping across a specimen depth of ~310 μm. When combined with expansion microscopy, cycleHCR revealed an intricate network of 10 subcellular structures in mouse embryonic fibroblasts. In mouse hippocampal slices, multiplex RNA and protein imaging uncovered complex gene expression gradients and cell-type-specific nuclear structural variations. cycleHCR provides a quantitative framework for elucidating spatial regulation in deep tissue contexts for research and has potential diagnostic applications.

Precision engineering of the probiotic Escherichia coli Nissle 1917 with prime editing.

CRISPR-Cas systems are transforming precision medicine with engineered probiotics as next-generation diagnostics and therapeutics. To promote human health and treat disease, engineering probiotic bacteria demands maximal versatility to enable non-natural functionalities while minimizing undesired genomic interferences. Here, we present a streamlined prime editing approach tailored for probiotic Escherichia coli Nissle 1917 utilizing only essential genetic modules, including Cas9 nickase from Streptococcus pyogenes, a codon-optimized reverse transcriptase, and a prime editing guide RNA, and an optimized workflow with longer induction. As a result, we achieved all types of prime editing in every individual round of experiments with efficiencies of 25.0%, 52.0%, and 66.7% for DNA deletion, insertion, and substitution, respectively. A comprehensive evaluation of off-target effects revealed a significant reduction in unintended mutations, particularly in comparison to two different base editing methods. Leveraging the prime editing system, we inserted a unique DNA sequence to barcode the edited strain and established an antibiotic-resistance-gene-free platform to enable non-natural functionalities. Our prime editing strategy presents a CRISPR-Cas system that can be readily implemented in any laboratories with the basic CRISPR setups, paving the way for future innovations in engineered probiotics.IMPORTANCEOne ultimate goal of gene editing is to introduce designed DNA variations at specific loci in living organisms with minimal unintended interferences in the genome. Achieving this goal is especially critical for creating engineered probiotics as living diagnostics and therapeutics to promote human health and treat diseases. In this endeavor, we report a customized prime editing system for precision engineering of probiotic Escherichia coli Nissle 1917. With such a system, we developed a barcoding system for tracking engineered strains, and we built an antibiotic-resistance-gene-free platform to enable non-natural functionalities. We provide not only a powerful gene editing approach for probiotic bacteria but also new insights into the advancement of innovative CRISPR-Cas systems.

Low-density SNP marker sets for genetic variation analysis and variety identification in cultivated citrus

BackgroundThe Citrus species are major fruit crops cultivated in the world and have complex genetic relationships due to sexual comparability between Citrus and related genera. Of these, satsuma mandarin (C. unshiu (Mak.) Marc.) and sweet orange (C. sinensis (L.) Osb.) are widely grown diploid species. In this study, genotyping by sequencing (GBS) was conducted to identify single nucleotide polymorphisms (SNPs) for investigating genetic variation in a citrus collection.ResultsA total of 26,903 high-quality SNPs were detected across nine chromosomes in the 144 citrus varieties, consisting of 70 C. unshiu, 40 C. sinensis, 22 interspecific hybrids, and 12 others. Of these, a core set of 481 SNPs was filtered based on polymorphism information content and genome distribution. Both principal component analysis (PCA) and model-based clustering showed genetic differentiation between C. unshiu and C. sinensis. For interspecific hybrids, these were separated from two species in PCA, but were mixed with each species in model-based clustering. Significant genetic differentiations between three populations were also found using the pairwise Fst. In addition, interspecific hybrids showed higher level of genetic diversity relative to the C. unshiu and C. sinensis populations. With the 481 SNPs, four subsets (192, 96, 48, and 24 SNPs) were generated to evaluate their performance for variety identification. Both 192 and 96 SNP sets distinguished all 144 varieties, while the 48 and 24 SNP sets separated 134 (93.1%) and 110 (76.4%), respectively.ConclusionsThe GBS-based SNP discovery led to robust and cost-effective molecular marker sets to assess genetic variation in the cultivated citrus species with narrow genetic bases. The resulting SNP sets are a resource to enhance the phenotype-based DUS testing by developing a DNA barcode system and thus facilitate new variety breeding and protection in citrus.

Implementation of radio frequency identification technology for a secure and intelligent shopping cart

Shopping at supermarkets has become a daily activity in urban areas of Enugu, Nigeria. However, there is always a huge rush in most mega supermarkets during times of discount offers, weekends and holidays resulting in long queues due to the barcode billing process. This research proposes a way of reducing the time spent at the billing counter using a radio frequency identification (RFID) smart-based shopping cart. To achieve this objective, an RFID tag, RFID reader, Arduino microcontroller and light-emitting diode (LED) display were used to develop a smart shopping trolley. RFID tag was placed on each of the eight products displayed for sale. RFID reader reads all the products that were placed on the cart and the details of the product such as the name, quantity, cost, and total cost was displayed on the LED. The smart shopping trolley system also incorporates an alarm system that triggers off when the RFID tag is removed from a product to avoid shoplifting and make the system secure for the owners of the supermarket. The result showed that the billing of the products was done directly from the smart shopping cart. The system was compared with the conventional barcode system and was found to overcome the limitations of time-consuming billing procedures.

Designing a Barcode System to Optimize Food Product Production and Distribution Management

Designing a Barcode System to Optimize Food Product Production and Distribution Management

Automatic medicine dispenser using medical prescriptions

In this paper, the objectives, the methodology used for the execution of the project and the results achieved is presented. The project constitutes the development of a self-service device that allows the delivery of medication prescribed by a specialist by reading an identifier on medical prescriptions. For development it was necessary different stages, the development of the barcode system in medical prescriptions was the first stage, considering that this system was carried out in a previous paper, subsequently the link was made with the electronic and programming part, that is, communication was established with the graphical user interface and subsequently the mechanical design for the integration of the device was generated. Adopting these tools will make it possible to achieve an important impact by reducing waiting times and preventing saturation of facilities in non-eventual situations, by contemplating their installation in different public places, expanding access for users to the health services network.

In Vivo Screening of Barcoded Gold Nanoparticles Elucidates the Influence of Shapes for Tumor Targeting

AbstractGold nanoparticles (NPs) are promising for cancer therapy due to their versatile shapes, optical properties, adjustable sizes, and facile functionalization. However, their diverse physicochemical properties, complex in vivo environments, and cellular heterogeneity lead to markedly different interactions with cells, impacting reproducibility and application selection. To address this, we implemented a DNA barcoding system to label gold NPs with varying shapes, sizes, and surface modifications for tumor delivery optimization. Six NP species, comprising three shapes (sphere, rod, triangle) and two sizes (40 nm, 80 nm), were tagged with unique DNA barcodes. Barcodes demonstrated minimal detachment, no interference with cell interactions, and resistance to DNase digestion. Next‐generation sequencing revealed that sphere NPs exhibited poor in vitro uptake but superior in vivo tumor targeting, likely due to enhanced endothelial interactions and reduced macrophage clearance. Additionally, 80 nm nanotriangles displayed excellent tumor targeting both in vitro and in vivo. Supporting experiments validated these findings. To demonstrate practical utility, the NPs were applied in siRNA delivery and photothermal therapy in a breast tumor model. This study pioneers the integration of DNA barcoding with gold NPs to systematically explore shape, size, and surface modifications for in vivo delivery.

Precise Lineage Tracking Using Molecular Barcodes Demonstrates Fitness Trade-offs for Ivermectin Resistance in Nematodes.

A fundamental tenet of evolutionary genetics is that the direction and strength of selection on individual loci varies with the environment. Barcoded evolutionary lineage tracking is a powerful approach for high-throughput measurement of selection within experimental evolution that to date has largely been restricted to studies within microbial systems, largely because the random integration of barcodes within animals is limited by physical and molecular protection of the germline. Here, we use the recently developed TARDIS barcoding system in Caenorhabditis elegans (Stevenson et al., 2023) to implement the first randomly inserted genomic-barcode experimental evolution animal model and use this system to precisely measure the influence of the concentration of the anthelmintic compound ivermectin on the strength of selection on an ivermectin resistance cassette. The combination of the trio of knockouts in neuronally expressed GluCl channels, avr-14, avr-15, and glc-1, has been previously demonstrated to provide resistance to ivermectin at high concentrations. Varying the concentration of ivermectin in liquid culture allows the strength of selection on these genes to be precisely controlled within populations of millions of individuals, yielding the largest animal experimental evolution study to date. The frequency of each barcode was determined at multiple time points via sequencing at deep coverage and then used to estimate the fitness of the individual lineages in the population. The mutations display a high cost to resistance at low concentrations, rapidly losing out to wildtype genotypes, but the balance tips in their favor when the ivermectin concentration exceeds 2nM. This trade-off in resistance is likely generated by a hindered rate of development in resistant individuals. Our results demonstrate that C. elegans can be used to generate high precision estimates of fitness using a high-throughput barcoding approach to yield novel insights into evolutionarily and economically important traits.

Overloading And unpacKing (OAK) - droplet-based combinatorial indexing for ultra-high throughput single-cell multiomic profiling

Multiomic profiling of single cells by sequencing is a powerful technique for investigating cellular diversity. Existing droplet-based microfluidic methods produce many cell-free droplets, underutilizing bead barcodes and reagents. Combinatorial indexing on microplates is more efficient for barcoding but labor-intensive. Here we present Overloading And unpacKing (OAK), which uses a droplet-based barcoding system for initial compartmentalization followed by a second aliquoting round to achieve combinatorial indexing. We demonstrate OAK’s versatility with single-cell RNA sequencing as well as paired single-nucleus RNA sequencing and accessible chromatin profiling. We further showcase OAK’s performance on complex samples, including differentiated bronchial epithelial cells and primary retinal tissue. Finally, we examine transcriptomic responses of over 400,000 melanoma cells to a RAF inhibitor, belvarafenib, discovering a rare resistant cell population (0.12%). OAK’s ultra-high throughput, broad compatibility, high sensitivity, and simplified procedures make it a powerful tool for large-scale molecular analysis, even for rare cells.

Chromosome-specific barcode system with centromeric repeat in cultivated soybean and wild progenitor.

Wild soybean Glycine soja is the progenitor of cultivated soybean Glycine max Information on soybean functional centromeres is limited despite extensive genome analysis. These species are an ideal model for studying centromere dynamics for domestication and breeding. We performed a detailed chromatin immunoprecipitation analysis using centromere-specific histone H3 protein to delineate two distinct centromeric DNA sequences with unusual repeating units with monomer sizes of 90-92 bp (CentGm-1) and 413-bp (CentGm-4) shorter and longer than standard nucleosomes. These two unrelated DNA sequences with no sequence similarity are part of functional centromeres in both species. Our results provide a comparison of centromere properties between a cultivated and a wild species under the effect of the same kinetochore protein. Possible sequence homogenization specific to each chromosome could highlight the mechanism for evolutionary conservation of centromeric properties independent of domestication and breeding. Moreover, a unique barcode system to track each chromosome is developed using CentGm-4 units. Our results with a unifying centromere composition model using CentGm-1 and CentGm-4 superfamilies could have far-reaching implications for comparative and evolutionary genome research.

Pembuatan Barcode Kemasan Produk Pada CV. Kuliner Makmur Sejahtera Kota Padang

Rendang, a traditional Minangkabau dish from West Sumatra, Indonesia, has gained international recognition and has inspired many entrepreneurs to promote this cuisine globally. One such entrepreneur is CV. Kuliner Makmur Sejahtera (KMS), which produces rendang under the brand Katuju. To enhance operational efficiency and support global market expansion, KMS requires an improved inventory management system. The primary challenge is the difficulty in monitoring inventory and tracking products without a barcode system. This community service program aims to address this issue by implementing barcode technology on Katuju's product packaging. The activities include designing, creating, and printing barcodes, as well as training KMS staff on how to use them. The process starts with observation and interviews to identify needs, followed by barcode design and production using an online barcode generator, and testing the barcode functionality with a scanning system. The results indicate that the community service team successfully trained KMS in barcode creation and printing, and integrated barcodes into product packaging. Evaluation through questionnaires showed participant satisfaction with the program and readiness to apply the barcode system to their products. This program is expected to improve inventory management, data accuracy, and support CV. KMS's global market expansion by providing tools necessary for efficient product tracking and valuable consumer data collection.

Fuzzy logic-based barcode scanning system for food products halal identification

Fuzzy logic-based barcode scanning system for food products halal identification

A cost-effective oligo-based barcode system for chromosome identification in longan and lychee.

Oligonucleotide (Oligo)-based fluorescence in situ hybridization (FISH) represents a highly effective methodology for identifying plant chromosomes. Longan is a commercially significant fruit species, yet lacking basic chromosomal markers has hindered its cytogenetic research. In this study, we developed a cost-effective oligo-based system for distinguishing chromosomes of longan (Dimocarpus longan Lour., 2n = 2x = 30). For this system, each synthesized oligo contained two chromosome-specific sequences that spanned a distance of over 200kb, and a PCR-based flexible amplification method coupled with nested primers was used for probe labeling. The use of these oligo-based barcodes enabled the marking of 36 chromosomal regions, which allowed for the unambiguous distinction of all 15 chromosomes in both longan and lychee (Litchi chinensis Sonn., 2n = 2x = 30) species. Based on the identification of individual chromosomes, we constructed karyotypes and detected genome assembly errors involving the 35S ribosomal RNA gene (35S rDNA) in longan and lychee. Developing oligo-based barcodes offers considerable promise for advancing cytogenetic research in longan, lychee, and their related species. Furthermore, this cost-effective synthesis system can be referred to the development of new oligo libraries among other species.

Development of Cost-Effective SNP Markers for Genetic Variation Analysis and Variety Identification in Cultivated Pears (Pyrus spp.).

Pear (Pyrus spp.) is a major fruit crop in the Rosaceae family, and extensive efforts have been undertaken to develop elite varieties. With advances in genome sequencing technologies, single-nucleotide polymorphisms (SNPs) are commonly used as DNA markers in crop species. In this study, a large-scale discovery of SNPs was conducted using genotyping by sequencing in a collection of 48 cultivated pear accessions. A total of 256,538 confident SNPs were found on 17 chromosomes, and 288 SNPs were filtered based on polymorphic information content, heterozygosity rate, and genome distribution. This subset of SNPs was used to genotype an additional 144 accessions, consisting of P. pyrifolia (53), P. ussuriensis (27), P. bretschneideri (19), P. communis (26), interspecific hybrids (14), and others (5). The 232 SNPs with reliable polymorphisms revealed genetic variations between and within species in the 192 pear accessions. The Asian species (P. pyrifolia, P. ussuriensis, and P. bretschneideri) and interspecific hybrids were genetically differentiated from the European species (P. communis). Furthermore, the P. pyrifolia population showed higher genetic diversity relative to the other populations. The 232 SNPs and four subsets (192, 96, 48, and 24 SNPs) were assessed for variety identification. The 192 SNP subset identified 173 (90.1%) of 192 accessions, which was comparable to 175 (91.1%) from the 232 SNPs. The other three subsets showed 81.8% (24 SNPs) to 87.5% (96 SNPs) identification rates. The resulting SNPs will be a useful resource to investigate genetic variations and develop an efficient DNA barcoding system for variety identification in cultivated pears.

Development of the Pneumococcal Genome Library, a core genome multilocus sequence typing scheme, and a taxonomic life identification number barcoding system to investigate and define pneumococcal population structure.

Investigating the genomic epidemiology of major bacterial pathogens is integral to understanding transmission, evolution, colonization, disease, antimicrobial resistance and vaccine impact. Furthermore, the recent accumulation of large numbers of whole genome sequences for many bacterial species enhances the development of robust genome-wide typing schemes to define the overall bacterial population structure and lineages within it. Using the previously published data, we developed the Pneumococcal Genome Library (PGL), a curated dataset of 30 976 genomes and contextual data for carriage and disease pneumococci recovered between 1916 and 2018 in 82 countries. We leveraged the size and diversity of the PGL to develop a core genome multilocus sequence typing (cgMLST) scheme comprised of 1222 loci. Finally, using multilevel single-linkage clustering, we stratified pneumococci into hierarchical clusters based on allelic similarity thresholds and defined these with a taxonomic life identification number (LIN) barcoding system. The PGL, cgMLST scheme and LIN barcodes represent a high-quality genomic resource and fine-scale clustering approaches for the analysis of pneumococcal populations, which support the genomic epidemiology and surveillance of this leading global pathogen.

Development of mitochondrial DNA cytochrome c oxidase subunit I primer sets to construct DNA barcoding library using next-generation sequencing.

Insects are one of the most diverse eukaryotic groups on the planet, with one million or more species present, including those yet undescribed. The DNA barcoding system has been developed, which has aided in the identification of cryptic species and undescribed species. The mitochondrial cytochrome c oxidase I region (mtDNA COI) has been utilised for the barcoding analysis of insect taxa. Thereafter, next-generation sequencing (NGS) technology has been developed, allowing for rapid acquisition of massive amounts of sequence data for genetic analyses. Although NGS-based PCR primers designed to amplify the mtDNA COI region have been developed, their target regions were only a part of COI region and/or there were taxonomic bias for PCR amplification. As the mtDNA COI region is a traditional DNA marker for the DNA barcoding system, modified primers for this region would greatly contribute to taxonomic studies. In this study, we redesigned previously developed PCR primer sets that targetted the mtDNA COI barcoding region to improve amplification efficiency and to enable us to conduct sequencing analysis on NGS. As a result, the redesigned primer sets achieved a high success rate (> 85%) for species examined in this study, covering four insect orders (Coleoptera, Lepidoptera, Orthoptera and Odonata). Thus, by combining the primers with developed primer sets for 12S or 16S rRNA regions, we can conduct more detailed taxonomic, phylogeographic and conservation genetic studies using NGS.

Offline Entry and Exit Gate Management System: A Barcode-Based Registration System at Jigme Namgyel Engineering College

The barcode system is used to automate data collection and is valuable to eliminate the possibility of human error, and the data obtained through the barcode is generated rapidly. Further, barcode scanning automatically enters a large amount of data into a system, making it efficient for recordkeeping and streamlining the records. Previously, Jigme Namgyel Engineering College (JNEC), had a paper-based registration system. It generated a great deal of paper, and time was consumed in long queues while students entered and exited the campus. Therefore, a QR system was developed whereby users scan outgoing/incoming QR through Google scanners and fill an online form. The system was also found to be time-consuming as the user had to punch in details manually and there was also a need for an internet connection. Although the system did help curb paper wastage. Therefore, to overcome the limitations of both systems, a Barcode System has been developed, wherein the barcode present in the user ID card is scanned and the user details will be auto-generated in the system. This system was developed and designed using Microsoft Access (MS Access). Problem-Based Learning (PBL) methodology was also implemented along with the System Development Life Cycle (SDLC). The system allows users to view and print the user record based on department. Generate users’ entry and exit reports, and also sort the report by date. The Barcode system is found to be the quickest and easiest way for the user to get registered. It is essential because it fulfills the primary role of keeping the record of outgoing or incoming users faster and easier. The Barcode system also fulfills the aim of going green by reducing paper wastage.