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Article: Designed gRNAs for CRISPR-Cas9 based antifungal resistance in eggplant.

Prajapati, Archana / Nain, Vikrant / Singh, Deepali

2023 Volume 19, Issue 8, Page(s) 844–848

Abstract: Eggplant is an important vegetable crop and is a good source of antioxidants, minerals, and vitamins. It has been used in ancient medicines for the treatment of multiple diseases. However, the cultivated varieties of eggplant are susceptible to numerous ... ...

Abstract	Eggplant is an important vegetable crop and is a good source of antioxidants, minerals, and vitamins. It has been used in ancient medicines for the treatment of multiple diseases. However, the cultivated varieties of eggplant are susceptible to numerous pathogens and pests that have a negative impact on vegetable crops. Increased resistance achieved through resistance genes (R genes) is limited in eggplant breeding due to the fact that R genes are typically specific to a pathogen race and can be quickly surpassed by pathogen evolution. The susceptibility genes (S genes) in plants facilitate pathogen entry and proliferation, thus disabling these genes might be beneficial for providing a broad range and durable resistance against pathogens. Reports on crops such as
Language	English
Publishing date	2023-08-31
Publishing country	Singapore
Document type	Journal Article
ZDB-ID	2203786-X
ISSN	0973-2063
ISSN	0973-2063
DOI	10.6026/97320630019844
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Article: Screening of CRISPR/Cas9 gRNA for mimicking Powdery Mildew resistant MLO ol-2 mutant.

Prajapati, Archana / Nain, Vikrant

Bioinformation

2021 Volume 17, Issue 6, Page(s) 637–645

Abstract: Powdery Mildew (PM) caused by fungal pathogen Oidium neolycopersici (O. neolycopersici) affects both greenhouse and field-grown tomato production. Resistance to PM disease can be achieved by selective inactivation of Mildew Resistance Locus O (MLO) genes ...

Abstract	Powdery Mildew (PM) caused by fungal pathogen Oidium neolycopersici (O. neolycopersici) affects both greenhouse and field-grown tomato production. Resistance to PM disease can be achieved by selective inactivation of Mildew Resistance Locus O (MLO) genes encoding heptahelical transmembrane domains, which confer susceptibility to fungal pathogens. Natural loss-of-function mutation is a 19 base pair (bp) deletion in the SlMLO1 gene locus responsible for fungal resistance in S. lycopersicum var. cerasiforme. Introgression of these resistance alleles through breeding into elite varieties is possible. However, this is a long and labour-intensive process and has limitations due to linkage drag. Nonetheless, recent developments in the field of genome editing technology particularly CRISPR/Cas9 systems allows quick, effective and accurate genome modification at the target gene locus. Therefore, it is of interest to determine the efficacy and exact deletion that mimics the natural ol-2 (Slmlo1) mutation present in wild tomatoes using CRISPR/Cas9. 947 putative guide RNAs (gRNAs) were designed using Cas9 variants to broaden Protospacer Adjacent Motif (PAM) compatibility and to enhance DNA specificity against the SlMLO1 locus. 60 out of 947 gRNAs were selected based on the recognition of the PAM sequence, the MIT specificity ranking, the off-target sites, their distance from the 19bp natural ol-2 mutation, the secondary structure of the gRNAs, and their minimum free energy. In depth analysis of these 60 gRNAs helped in the selection of the top five gRNAs based on the above-mentioned criteria. These gRNAs are useful for introducing deletions identical to natural ol-2 mutants and impart resistance against fungal pathogen O. neolycopersici in cultivated tomato crops.
Language	English
Publishing date	2021-06-30
Publishing country	Singapore
Document type	Journal Article
ZDB-ID	2203786-X
ISSN	0973-2063
ISSN	0973-2063
DOI	10.6026/97320630017637
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Article ; Online: TALENs-an indispensable tool in the era of CRISPR: a mini review.

Bhardwaj, Anuradha / Nain, Vikrant

Journal, genetic engineering & biotechnology

2021 Volume 19, Issue 1, Page(s) 125

Abstract: Background: Genome of an organism has always fascinated life scientists. With the discovery of restriction endonucleases, scientists were able to make targeted manipulations (knockouts) in any gene sequence of any organism, by the technique popularly ... ...

Abstract	Background: Genome of an organism has always fascinated life scientists. With the discovery of restriction endonucleases, scientists were able to make targeted manipulations (knockouts) in any gene sequence of any organism, by the technique popularly known as genome engineering. Though there is a range of genome editing tools, but this era of genome editing is dominated by the CRISPR/Cas9 tool due to its ease of design and handling. But, when it comes to clinical applications, CRISPR is not usually preferred. In this review, we will elaborate on the structural and functional role of designer nucleases with emphasis on TALENs and CRISPR/Cas9 genome editing system. We will also present the unique features of TALENs and limitations of CRISPRs which makes TALENs a better genome editing tool than CRISPRs. Main body: Genome editing is a robust technology used to make target specific DNA modifications in the genome of any organism. With the discovery of robust programmable endonucleases-based designer gene manipulating tools such as meganucleases (MN), zinc-finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and clustered regularly interspaced short palindromic repeats associated protein (CRISPR/Cas9), the research in this field has experienced a tremendous acceleration giving rise to a modern era of genome editing with better precision and specificity. Though, CRISPR-Cas9 platform has successfully gained more attention in the scientific world, TALENs and ZFNs are unique in their own ways. Apart from high-specificity, TALENs are proven to target the mitochondrial DNA (mito-TALEN), where gRNA of CRISPR is difficult to import. This review talks about genome editing goals fulfilled by TALENs and drawbacks of CRISPRs. Conclusions: This review provides significant insights into the pros and cons of the two most popular genome editing tools TALENs and CRISPRs. This mini review suggests that, TALENs provides novel opportunities in the field of therapeutics being highly specific and sensitive toward DNA modifications. In this article, we will briefly explore the special features of TALENs that makes this tool indispensable in the field of synthetic biology. This mini review provides great perspective in providing true guidance to the researchers working in the field of trait improvement via genome editing.
Language	English
Publishing date	2021-08-21
Publishing country	Netherlands
Document type	Journal Article ; Review
ZDB-ID	2637420-1
ISSN	2090-5920 ; 1687-157X ; 2090-5920
ISSN (online)	2090-5920
ISSN	1687-157X ; 2090-5920
DOI	10.1186/s43141-021-00225-z
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Article: Towards the diagnosis of dengue virus and its serotypes using designed CRISPR/Cas13 gRNAs.

Prajapati, Archana / Tandon, Ashmita / Nain, Vikrant

Bioinformation

2022 Volume 18, Issue 8, Page(s) 661–668

Abstract: Dengue Virus (DENV) is a mosquito-borne virus that is prevalent in the world's tropical and subtropical regions. Therefore, early detection and surveillance can help in the management of this disease. Current diagnostic methods rely primarily on ELISA, ... ...

Abstract	Dengue Virus (DENV) is a mosquito-borne virus that is prevalent in the world's tropical and subtropical regions. Therefore, early detection and surveillance can help in the management of this disease. Current diagnostic methods rely primarily on ELISA, PCR, and RT-PCR, among others, which can only be performed in specialized laboratories and require sophisticated instruments and technical expertise. CRISPR-based technologies on the other hand have field-deployable viral diagnostics capabilities that could be used in the development of point-of-care molecular diagnostics. The first step in the field of CRISPR-based virus diagnosis is to design and screen gRNAs for high efficiency and specificity. In the present study, we employed a bioinformatics approach to design and screen DENV CRISPR/Cas13 gRNAs for conserved and serotype-specific variable genomic regions in the DENV genome. We identified one gRNA sequence specific for each of the lncRNA and NS5 regions and identified one gRNA against each of DENV1, DENV2, DENV3, and DENV4 to distinguish the four DENV serotypes. These CRISPR/Cas13 gRNA sequences will be useful in diagnosing the dengue virus and its serotypes for in vitro validation and diagnostics.
Language	English
Publishing date	2022-08-31
Publishing country	Singapore
Document type	Journal Article
ZDB-ID	2203786-X
ISSN	0973-2063
ISSN	0973-2063
DOI	10.6026/97320630018661
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Article ; Online: In silico

Suman / Chaudhary, Meenakshi / Nain, Vikrant

Journal of biomolecular structure & dynamics

2020 Volume 39, Issue 3, Page(s) 841–850

Abstract: Cold shock domain (CSD) proteins with nucleic acid binding properties are well conserved from bacteria to higher organisms. In bacteria, the cold shock proteins (CSPs) are single domain RNA chaperones, whereas in animals and plants, CSDs are accompanied ... ...

Abstract	Cold shock domain (CSD) proteins with nucleic acid binding properties are well conserved from bacteria to higher organisms. In bacteria, the cold shock proteins (CSPs) are single domain RNA chaperones, whereas in animals and plants, CSDs are accompanied by additional domains with roles in transcription regulation. Bacterial CSPs (
MeSH term(s)	Bacillus subtilis/genetics ; Bacterial Proteins/genetics ; Base Sequence ; Cold Shock Proteins and Peptides/genetics ; Cold Temperature ; Cold-Shock Response ; Molecular Docking Simulation ; Phylogeny ; RNA, Plant
Chemical Substances	Bacterial Proteins ; Cold Shock Proteins and Peptides ; RNA, Plant
Language	English
Publishing date	2020-02-12
Publishing country	England
Document type	Journal Article
ZDB-ID	49157-3
ISSN	1538-0254 ; 0739-1102
ISSN (online)	1538-0254
ISSN	0739-1102
DOI	10.1080/07391102.2020.1719198
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Article ; Online: Rapamycin and Torin2 inhibit

Biswas, Biswambhar / Gangwar, Garima / Nain, Vikrant / Gupta, Ishaan / Thakur, Anil / Puria, Rekha

Journal of biomolecular structure & dynamics

2022 Volume 41, Issue 17, Page(s) 8445–8461

Abstract: ... The ... ...

Abstract	The fungus
Language	English
Publishing date	2022-10-20
Publishing country	England
Document type	Journal Article
ZDB-ID	49157-3
ISSN	1538-0254 ; 0739-1102
ISSN (online)	1538-0254
ISSN	0739-1102
DOI	10.1080/07391102.2022.2134927
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Article ; Online: VEGF-mediated tumour growth and EMT in 2D and 3D cell culture models of hepatocellular carcinoma.

Rawal, Preety / Tripathi, Dinesh Mani / Nain, Vikrant / Kaur, Savneet

Oncology letters

2022 Volume 24, Issue 3, Page(s) 315

Abstract: The purpose of the present study was to evaluate the effects of vascular endothelial growth factor (VEGF) on tumorigenic properties in two-dimensional (2D) and three-dimensional (3D) cultures of hepatoma cells. The proliferation and invasion of hepatoma ... ...

Abstract	The purpose of the present study was to evaluate the effects of vascular endothelial growth factor (VEGF) on tumorigenic properties in two-dimensional (2D) and three-dimensional (3D) cultures of hepatoma cells. The proliferation and invasion of hepatoma cells was assessed using wound healing, chemotaxis Transwell, invasion, tube-forming and hanging drop assays in both 2D and 3D cultures. The expression levels of epithelial-mesenchymal transition (EMT) and stemness markers were analysed using reverse transcription-quantitative PCR (RT-qPCR) for mRNA expression and immunofluorescence assay for protein expression. To validate the role of VEGF in tumour growth, a VEGF receptor (VEGFR) inhibitor (sorafenib) was used. The results demonstrated that the hepatoma cells formed 3D spheroids that differed in size and density in the absence and presence of the growth factor, VEGF. In all spheroids, invasion and angiogenesis were more aggressive in 3D cultures in comparison to 2D conditions following treatment with VEGF. Mechanistically, the VEGF-mediated increase in the levels of EMT markers, including Vimentin, N-cadherin 2 (Cadherin 2) and Thy-1 Cell Surface Antigen was observed in the 2D and 3D cultures. Sorafenib treatment for 24 h culminated in a marked reduction in cell migration, cell-cell adhesion, spheroid compaction and EMT gene expression in 3D models as compared to the 2D models. On the whole, the findings of the present study suggested that as compared to the 2D cell cultures, 3D cell cultures model may be used as a more realistic model for the study of tumour growth and invasion in the presence of angiogenic factors, as well as for tumour inhibitor screening.
Language	English
Publishing date	2022-07-15
Publishing country	Greece
Document type	Journal Article
ZDB-ID	2573196-8
ISSN	1792-1082 ; 1792-1074
ISSN (online)	1792-1082
ISSN	1792-1074
DOI	10.3892/ol.2022.13435
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Article ; Online: A convenient and robust protocol for preparation of ODAP-free Lathyrus sativus protein.

Suman / Ahmad, Yasmin / Nain, Vikrant

Analytical biochemistry

2019 Volume 591, Page(s) 113544

Abstract: The present study aimed to develop a protocol for easy removal of β-ODAP neurotoxin by converting it into its isomer α-ODAP (reported to be less toxic) followed by its separation from the protein fraction in pH dependent manner. Use of β-mercaptoethanol ... ...

Abstract	The present study aimed to develop a protocol for easy removal of β-ODAP neurotoxin by converting it into its isomer α-ODAP (reported to be less toxic) followed by its separation from the protein fraction in pH dependent manner. Use of β-mercaptoethanol prevented aggregate formation and increased solubility of the prepared Lathyrus sativus protein. Validation of ODAP removal by paper chromatography and mass spectrometry indicated the robustness of the protocol. Removal of ODAP and presence of high antioxidants and homoarginine content can enable Lathyrus sativus to be an alternate source of protein, as well as have other health benefits, including benefits for patients with cardiovascular diseases.
MeSH term(s)	Amino Acids, Diamino/isolation & purification ; Isomerism ; Lathyrus/chemistry ; Plant Extracts/chemistry ; Plant Proteins/isolation & purification ; Seeds/chemistry
Chemical Substances	Amino Acids, Diamino ; Plant Extracts ; Plant Proteins ; oxalyldiaminopropionic acid (1TG777QI25)
Language	English
Publishing date	2019-12-19
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	1110-1
ISSN	1096-0309 ; 0003-2697
ISSN (online)	1096-0309
ISSN	0003-2697
DOI	10.1016/j.ab.2019.113544
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Article ; Online: Molecular docking and dynamic simulation analysis of

Chaudhary, Meenakshi / Nain, Vikrant / Sehgal, Deepak

Journal of biomolecular structure & dynamics

2021 Volume 41, Issue 4, Page(s) 1342–1350

Abstract: The unavailability of a suitable treatment for human Hepatitis E virus (HEV) infection necessitate the development of anti HEV drugs. The HEV papain-like cysteine proteases (HEV PCP) is a crucial target to prevent viral replication and progression. E64 ... ...

Abstract	The unavailability of a suitable treatment for human Hepatitis E virus (HEV) infection necessitate the development of anti HEV drugs. The HEV papain-like cysteine proteases (HEV PCP) is a crucial target to prevent viral replication and progression. E64 is a known HEV PCP inhibitor; however, its molecular mechanism of inhibition is not yet known. Since the crystal structure of HEV PCP is not available, the primary focuses of the present study was to refine the predicted HEV PCP structural model by molecular dynamics (MD) simulation. Further, we performed a 200 ns MD simulation to understand the structural complexity of HEV PCP and the effect of E64 binding with HEV PCP. The E64 binding with active site residues Gln48, Thr51, Gln55, Cys52, Ser81, Gln 98, Cys 132, Arg158, His159, Asn 160 and Ala96 leads to reduced fluctuations in the residue at N-terminal (18-41) that include the CHC motif (26-28). However, most of the other non interacting residues, including the inter-domain linker region (46-87), showed increased fluctuations in the HEV PCP-E64 complex. The residue Asp21 and Ala96 are involved in the formation of interdomain interactions in the HEV PCP apo enzyme. While in the PCP-E64 complex, E64 binds to Ala96 and creates a steric hindrance to prevent interdomain interactions. Thus, the E64 binding reduces interdomain interactions and restrict domain movements in the HEV PCP-E64 complex. This information will be important for the chemically designing more effective derivatives of E64 developing HEV PCP specific inhibitors.Communicated by Ramaswamy H. Sarma.
MeSH term(s)	Humans ; Catalytic Domain ; Endopeptidases ; Hepatitis E virus/enzymology ; Hepatitis E virus/physiology ; Molecular Docking Simulation ; Molecular Dynamics Simulation ; Peptide Hydrolases ; Viral Papain-like Proteases/antagonists & inhibitors ; Viral Papain-like Proteases/metabolism
Chemical Substances	Endopeptidases (EC 3.4.-) ; Peptide Hydrolases (EC 3.4.-) ; Viral Papain-like Proteases (EC 3.4.22.-)
Language	English
Publishing date	2021-12-21
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	49157-3
ISSN	1538-0254 ; 0739-1102
ISSN (online)	1538-0254
ISSN	0739-1102
DOI	10.1080/07391102.2021.2019124
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Article: A convenient and robust protocol for preparation of ODAP-free Lathyrus sativus protein

Suman / Ahmad, Yasmin / Nain, Vikrant

Analytical biochemistry. 2020 Feb. 15, v. 591

2020

Abstract	The present study aimed to develop a protocol for easy removal of β-ODAP neurotoxin by converting it into its isomer α-ODAP (reported to be less toxic) followed by its separation from the protein fraction in pH dependent manner. Use of β-mercaptoethanol prevented aggregate formation and increased solubility of the prepared Lathyrus sativus protein. Validation of ODAP removal by paper chromatography and mass spectrometry indicated the robustness of the protocol. Removal of ODAP and presence of high antioxidants and homoarginine content can enable Lathyrus sativus to be an alternate source of protein, as well as have other health benefits, including benefits for patients with cardiovascular diseases.
Keywords	Lathyrus sativus ; antioxidants ; beta-mercaptoethanol ; cardiovascular diseases ; isomers ; mass spectrometry ; neurotoxins ; pH ; paper chromatography ; patients ; solubility ; toxicity
Language	English
Dates of publication	2020-0215
Publishing place	Elsevier Inc.
Document type	Article
ZDB-ID	1110-1
ISSN	1096-0309 ; 0003-2697
ISSN (online)	1096-0309
ISSN	0003-2697
DOI	10.1016/j.ab.2019.113544
Database	NAL-Catalogue (AGRICOLA)

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