Research Article
Engineering Probiotic Strains for Gut Health Enhancement Using CRISPR and Molecular Marker-assisted Technologies
Issue:
Volume 14, Issue 1, June 2026
Pages:
1-12
Received:
18 December 2025
Accepted:
29 December 2025
Published:
19 January 2026
Abstract: Probiotics play a vital role in maintaining gut homeostasis, modulating immune responses, and promoting overall human health. Traditional approaches to probiotic strain development rely primarily on natural isolation and phenotypic screening, which are time-consuming and lack precision. The current study presents an in silico bioinformatics framework for the rational enhancement of probiotic strains through CRISPR-Cas9–guided design, integrated with structural bioinformatics and immunoinformatics analyses. Sequence homology and conservation were evaluated using BLAST and multiple sequence alignment to identify suitable genetic targets while minimizing off-target similarity. Structural insights were obtained from the PDB and MMDB, with PDB ID 2Z7X which was a representative immune-related protein model. Structural stability and conformational variation of hypothetical modifications were assessed using RMSD-based comparisons. Guide RNA candidates for genome editing were computationally nominated and ranked using E-CRISP and CHOPCHOP, emphasizing predicted efficiency and reduced off-target risk. To evaluate immunological safety, reverse vaccinology–based B-cell epitope prediction was performed using BepiPred, with epitope regions mapped onto three-dimensional protein structures. The integrated pipeline enables the identification of modification-tolerant regions while minimizing immunogenic potential. This purely computational strategy reduces experimental dependency, accelerates strain optimization, and provides a reproducible foundation for future probiotic engineering studies under appropriate biosafety and regulatory frameworks. This study provides a computational foundation for designing safer and more effective probiotic strains for gut health, immunomodulation, and disease prevention. The framework can support functional food development, precision microbiome therapies, vaccine-adjuvant research, and regulatory pre-screening of engineered probiotics while minimizing laboratory costs and biosafety risks.
Abstract: Probiotics play a vital role in maintaining gut homeostasis, modulating immune responses, and promoting overall human health. Traditional approaches to probiotic strain development rely primarily on natural isolation and phenotypic screening, which are time-consuming and lack precision. The current study presents an in silico bioinformatics framewo...
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Research Article
Structure-guided CRISPR CAS9 Targeting of ABL1 for Functional Disruption of BCR-ABL1 Fusion in Chronic Myeloid Leukaemia
Issue:
Volume 14, Issue 1, June 2026
Pages:
13-25
Received:
10 January 2026
Accepted:
21 January 2026
Published:
4 February 2026
Abstract: Chronic myeloid leukaemia (CML) is a clonal myeloproliferative cancer caused by the constant activity of the BCR–ABL 1 fusion protein' s tyrosine kinase, resulting from the Philadelphia chromosome translocation, which leads to abnormal cell growth, survival, and disease progression. While tyrosine kinase inhibitors (TKIs) have greatly improved patient outcomes, issues like drug resistance and persistent leukemic stem cells highlight the need for alternative therapies. This study used a structure- guided CRISPR- Cas 9 genome editing approach to identify highly specific single- guide RNAs (sgRNAs) that can disrupt the human ABL 1 gene, a key part of the BCR–ABL 1 fusion. The high- resolution crystal structure of the ABL 1 kinase domain (PDB ID: 8 I 7 S) helped identify essential functional regions, including catalytic and ATP- binding sites, for precise CRISPR targeting. Computational design and filtering of sgrnas were performed using E- CRISP and CHOPCHOP, focusing on criteria like PAM site accessibility, targeting conserved kinase regions in exons, GC content, predicted efficiency, and low off- target risk. In silico analyses, including specificity scores, mismatch profiles, and sequence alignment across ABL 1 transcript variants, ensured high selectivity and broad coverage. Genomic visualization confirmed accurate targeting within exons encoding vital kinase functions. Protein–protein interaction analysis via STRING showed strong links between ABL 1 and key oncogenic regulators such as BCR, STAT 5, and MAPK pathway components. KEGG pathway analysis further indicated ABL 1' s involvement in chronic myeloid leukaemia, PI 3 K–AKT, MAPK signaling, and other cancer- related pathways, emphasising its importance in CML development. This combined computational approach demonstrates that structure- guided CRISPR- Cas 9 targeting of ABL 1 can effectively disrupt BCR–ABL 1 driven cancer signals. The results provide a strong theoretical basis for future experimental validation and genome editing therapies aimed at overcoming TKI resistance and achieving long- lasting molecular remission in CML.
Abstract: Chronic myeloid leukaemia (CML) is a clonal myeloproliferative cancer caused by the constant activity of the BCR–ABL 1 fusion protein' s tyrosine kinase, resulting from the Philadelphia chromosome translocation, which leads to abnormal cell growth, survival, and disease progression. While tyrosine kinase inhibitors (TKIs) have greatly improved pati...
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Research Article
Integrative GWAS and CRISPR Functional Validation of Gene Variant Structure of AtHKT1; 1 in KCl
Shephali Sachan
,
Uma Kumari*
Issue:
Volume 14, Issue 1, June 2026
Pages:
26-40
Received:
1 April 2026
Accepted:
11 April 2026
Published:
25 April 2026
DOI:
10.11648/j.cbb.20261401.13
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Abstract: Understanding ion transport mechanisms in plants is crucial for enhancing stress tolerance and crop yields. The Arabidopsis thaliana High-Affinity Potassium Transporter 1; 1 (AtHKT1; 1) is vital for maintaining sodium and potassium balance under saline stress. However, the structural and functional effects of genetic variants in AtHKT1; 1, especially in potassium chloride (KCl) environments, are not fully understood. This research combiness Genome-Wide Association Studies (GWAS) with CRISPR-based functional validation to examine AtHKT1; 1 gene variants, focusing on the protein structure with PDB ID: 8W9O. The study first used GWAS and SNP discovery to find significant genetic variations linked to ion transport and salt tolerance. Candidate SNPs were selected based on their statistical significance and potential biological roles. Structural analysis of the protein 8W9O involved PDB and MMDB resources, with validation through ERRAT and visualization/mutation mapping in PyMOL. InterProScan was used to identify conserved functional motifs. To validate SNP effects, CRISPR guide RNAs were designed with E-CRISP and CHOPCHOP, targeting key gene regions for precise editing. This integrated approach linked genetic variations to structural changes and their potential impact on ion binding and transport, especially under KCl conditions. Results showed certain SNPs cause conformational shifts in key transmembrane regions of AtHKT1; 1, possibly influencing ion selectivity and transport efficiency. Structural validation confirmed the accuracy of the modeled variants, and domain analysis revealed disruptions in conserved functional areas. CRISPR strategies proved feasible for precise gene editing to test these functional hypotheses. Overall, this study offers a comprehensive framework that combines GWAS, structural bioinformatics, and CRISPR technology to explore how genetic variants affect AtHKT1; 1 function. These insights improve understanding of ion transport regulation in saline environments and support the development of genetically modified crops with enhanced salt tolerance.
Abstract: Understanding ion transport mechanisms in plants is crucial for enhancing stress tolerance and crop yields. The Arabidopsis thaliana High-Affinity Potassium Transporter 1; 1 (AtHKT1; 1) is vital for maintaining sodium and potassium balance under saline stress. However, the structural and functional effects of genetic variants in AtHKT1; 1, especial...
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