http://nopr.niscpr.res.in/handle/123456789/58532 2026-10-11T13:09:11Z 2026-10-11T13:09:11Z Palan, Bhavesh Bhattacharya, Anjanabha Char, Bharat http://nopr.niscpr.res.in/handle/123456789/58542 2021-11-17T11:38:15Z 2021-01-01T00:00:00Z

Title: TILLING in the era of precise genome editing Authors: Palan, Bhavesh; Bhattacharya, Anjanabha; Char, Bharat Abstract: To feed the ceaselessly growing population, it is challenging to increase the food productivity with limited land and water resources under changing climatic regimes. Conventional breeding has played vital role in increasing the crop productivity however takes long time and is labor intensive. Later, the transgenic technology came in to existence and played an important role for increased crop production. However, due to the regulatory hurdles in several countries including India, restricted extensive commercialization of genetically modified (GM) crops was possible. Therefore, more emphasis is now needed towards non-GM techniques for crop improvement. Now-a-days, genome editing (GE) techniques are also employed in crop improvement projects because of its simplicity, robustness and high efficiency. Parallel to GE, TILLING (Targeting Induced Local Lesions IN Genome) which is a type of random GE can be used as a non-transgenic approach, which involves developing a large mutant population in wide range of crops and screening the population for mutants. Once a mutant population is produced, it can be TILLed any number of times. Therefore, TILLING is a rapid, simple, low cost, effective, high-throughput and is independent of genotype and genome size. In this review, we focused on the potential and application of TILLING technique in crop improvement programs, in the era of precise genome editing. Page(s): 9-16

2021-01-01T00:00:00Z Khoiri, Syaiful Giyanto Damayanti, Tri Asmira http://nopr.niscpr.res.in/handle/123456789/58541 2021-11-17T11:34:22Z 2021-01-01T00:00:00Z

Title: Characterization of acyl-homoserine lactonase gene from Brevibacillus brevis strain B37 Authors: Khoiri, Syaiful; Giyanto; Damayanti, Tri Asmira Abstract: Acyl-homoserine lactonase (EC 3.1.1.25) is a metallo-betalactamase, specifically hydrolyzed N-acyl-homoserine lactones (AHL) secreted by Gram-negative bacteria. AHL lactonase has been reported as a potential substitute for synthetic anti-bacterial, such as reduce the severity of plant diseases caused by Xanthomonas campestris pv. campestris, and Pectobacterium catrotovorum. The exploration of lactonase producing organisms has been widely reported. AHL-lactonase is produced by Bacillaceae bacteria such as Bacillus thuringiensis, B. cereus, and B. antrachis. AHL-lactonase produced by Bacillaceae bacteria was translated from aiiA gene. In our previous study, aiiA novel gene was detected in Brevibacillus brevis B37 but has not been characterized. This study aimed to clone aiiA gene isolated from B.brevis B37 by polymerase chain reaction (PCR) with a pair of degenerated primers, to reveal homology comparison withothers aiiA genes and amino acids, to express aiiA gene in Escherichia coli BL21 (DE3), and also to assay quorum quencherability. The aiiA gene was successfully isolated with 753 bp and 250 amino acids. The aiiA gene and the AiiA protein fromB.brevis B37 had high similarity with aiiA and AiiA from B. thuringiensis group. The deduced amino acid sequencecontained conserved sequence region 103SHLHFDH109 and 166TPGHTPGH173 as characteristic of the metallo betalactamasefamily. Additionally, the aiiAB37 gene was expressed in E. coli BL21 (DE3) and the expressed AiiA protein could attenuate the expression of violacein produce by Chromobacterium violaceum and decrease the expression of soft rot symptom caused by Dickeya dadantii. Page(s): 17-25

2021-01-01T00:00:00Z Sarkar, Swaroop Suryanarayana, V V S Reddy, G R Dechamma, H J Shankar, S R Madhan http://nopr.niscpr.res.in/handle/123456789/58540 2021-11-17T11:29:58Z 2021-01-01T00:00:00Z

Title: Creation of leaderless FMDV replicon for development of replication defective virus (leaderless FMDV): A strategy towards the development of attenuated vaccine with marker facility Authors: Sarkar, Swaroop; Suryanarayana, V V S; Reddy, G R; Dechamma, H J; Shankar, S R Madhan Abstract: Leader protease (Lpro) of foot-and-mouth disease virus (FMDV) which is essential for viral replication and pathogenicity in host is found in two forms, Lab and Lb with similar activity but, both differing only at amino-termini with separate initiation codons, 84 nucleotides apart. After translation of the genomic RNA into single polyprotein, the L protease is released autocatalytic from the N terminus and cleaves the p220 subunit of the eukaryotic initiation factor 4F complex resulting in the shut off host protein synthesis, an essential step for viral pathogenicity. We exploited this function for development of attenuated virus by deleting the gene encoding Lb protease from FMDV Asia 1(63/72) cDNA replicon (Joshi et al, 2013) by PCR mutagenesis approach. The deletions and intactness of the frames were confirmed by sequence analysis. P1-2A polyprotein gene of FMDV ‘O’ was inserted into the replicon and the full length construct was studied for virulence to baby hamster kidney (BHK) 21 cells. Vaccinia expressing T7 polymerase was used for in vitro RNA generation and infection. The lower cytopathic effect (cpe) as observed by reduced replication efficiency confirmed the effect of Lb deletion when compared with the construct with no deletion. Page(s): 26-34

2021-01-01T00:00:00Z Sarkar, Swaroop Suryanarayana, V V S Sekar, S Chandra Shankar, S R Madan Reddy, G R Dechamma, H J http://nopr.niscpr.res.in/handle/123456789/58539 2021-11-17T11:22:30Z 2021-01-01T00:00:00Z

Title: A strategy to develop disabled infectious single-cycle (DISC) foot and mouth disease virus by 3B3 gene deletion using the infective cDNA copy of the genome Authors: Sarkar, Swaroop; Suryanarayana, V V S; Sekar, S Chandra; Shankar, S R Madan; Reddy, G R; Dechamma, H J Abstract: Disabled infectious single-cycle (DISC) virus is in between attenuated and inactivated. When used as a vaccine DISC virus behaves like inactivated virus as it cannot further multiply in the vaccinated individual after one cycle of replication. When infected to permissive cells expressing virus specific protein which has been deleted from viral genome, the virus replicates normally. The development of DISC virus involves the deletion of an open reading frame (ORF) coding for a key protein involved in the viral replication or viral capsid formation. Such virus, when injected in animals, can complete only one round of replication without producing a progeny virus. Here we report similar strategy followed for the production DISC foot and mouth disease virus (FMDV). We have selected 3B3 protein gene that expresses 3 copies of virus specific genome-linked protein needed for virus replication and deleted from the FMDV replicon carrying FMDV serotype Asia 1 backbone and inserted the same into baby hamster kidney 21 (BHK-21) cell genome. Upon transfection of the genetically modified BHK cells with RNA copy of the genetically modified cDNA, replication of the virus started in these cells. The DISC virus so developed can be a potent vaccine candidate for achieving robust and long duration of immune response against FMDV infection in bovine. The approach also took care in the development of serotype specific DISC viruses using single FMDV Asia 1 replicon. Vaccines based on DISC viruses may be superior in terms of immune response as compared to inactivated vaccines. Page(s): 35-42

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