Dr. Nirmalya Ganguli did his Bachelor’s and Master’s Degree from Bangalore University. He obtained his Ph.D. degree from the University of Delhi South Campus, and his doctoral work was focused on Direct Delivery of Transgene in the Mammary Gland for production of Therapeutic Protein. He worked as Research Associate at the National Institute of Immunology, New Delhi, on developing easy methods of transgenesis through testicular route as well as Spermatogonial Stem Cells/Germ Cell transplantation study. He joined NIAB in the year 2017.
Dr. Ganguli is working on development of genetically engineered livestock for agricultural applications and biomedical research. He has specialization in the area of Transgenesis, Generation of Animal Bioreactor, Direct Mammary Gland Transfection, Genome and Protein Engineering. He is experienced in the generation of transgenic mice, generation of gene knockdown mice using shRNA, genome engineering through CRISPR-Cas system, generation of various ubiquitous/tissue specific mammalian expression vector, production of therapeutic protein in milk, and testicular germ cell depletion & stem cell transplantation work.
Dr. Ganguli’s group is presently working for development of easy method of transgenesis in farm animals. This will help in generation of farm animal models of diseases and development of disease resistance and/or high milk yielding animals. Along with the lab of Dr. Subeer S. Majumdar, work is also being carried on development of non-GMO animal bioreactor by directly transfecting the epithelial cells of udder gland for production of animal/human therapeutic proteins in milk. His group is standardising the testicular germ cell depletion and spermatogonial stem cell transplantation technology in farm animals to serve various purposes. This will also help in production of transgenic sperm by specific and precise genomic modification (site specific integration or deletion). Besides this work is also going on to adopt CRISPR/Cas technology for precise genome editing in farm animals for increasing productivity. Along with this, his group also intends to investigate the cellular signaling involved during udder gland development in farm animals which may help in modifying components of milk.
Establishing Testicular Transgenesis in Farm Animals:
The laboratory has expertise in generation of transgenic animals through testicular transgenesis. This involves electroporation or hypotonic shock mediated delivery of transgene into germ cells, which eventually develops into sperm. Such sperm are used to generate transgenic animals. This procedure is well studied in mice and successfully generated transgene bearing pups in a litter. The Lab at present working towards adopting this simple and less cumbersome technique for generation of transgenic farm animals as the conventional methods of generation of Transgenic farm animals are cumbersome, time consuming and require huge infrastructure.
Figure: A: Shows the in-vivo testicular electroporation in Rabbit testis. B: Shows the expression of EGFP in the cultured testicular tubule of rabbit, after electroporation with EGFP expressing transgene construct. C: Shows the expression of EGFP in the cultured testicular tubule of goat, after electroporation with EGFP expressing transgene construct.
We are also working towards enhancing the efficiency of the testicular transgenesis for farm animals. We are performing Slot-Blot and Droplet Digital PCR to check integration of the transgene followed by multiple fluorophores based Confocal Microscopy to detect subcellular localization of the expressed exogenous protein.
Figure: We are fusing/engineering signal peptides of different sperm membrane glycol-protein to translocate exogenous protein (like EGFP) on the sperm membrane. A. Confocal microscopy image shows immuno cytochemistry analysis of HEK293 cells expressing EGFP fused with transmembrane domain of sperm surface glycol protein (mFused-EGFP) along with construct pColourful-Cell (Addgene plasmid # 62449). EGFP is localized on the cell membrane as confirmed by co localization of membrane tagged Cerulean. B. Image shows 2.5D analysis for estimating the Fluorescence Intensity of mFused-EGFP(counter stained with A546) Transfected Permeabilized and Non Permeabilized HEK293 Cells using Zen 2.1 software from Zeiss. Difference in Fluorescence intensity in non-permeabilised cells as compared to permeabilized cells confirm the presence of mFused-EGFP on the surface of the cell membrane. C. Shows the magnified view of the area marked in the image A.
We have developed expertise in generating multi-cistronic mammalian expression vector through synthetic biology approach for ubiquitous and cell/tissue specific expression of desired gene through transgenesis. All together this will also help in easy and fast generation of transgenic farm animal bioreactor for high yielding and/or disease resistant animal herd as well as models for various animal diseases.
Generation of Animal Bioreactor for Production of Biotherapeutics:
With increasing knowledge about functions of genes/gene products (proteins), relation of their deficiencies with causation of a particular abnormal condition of the body is getting divulged. For their replenishment, we need to produce these proteins in bulk amount as a therapeutic agent (biotherapeutics), so that this knowledge is translated into therapy and diseases remediation. In the present scenario, production of Biopharmaceuticals at cheaper cost is the need of the world. Developing better bioprocessing technologies and overcoming weak development pipelines may help in achieving this objective. We are working with Dr. Majumdar to develop an easy and cost-effective alternative approach to generate mammary gland bioreactor for production of various bio therapeutics by passing the GMO related issue. Instead of many existing methods, generation of fast non-GMO animal bioreactor remains a challenge till date. We have established virosome based direct transfection of transgene in mammary epithelial cells followed by production of biotherapeutics in the milk in mice and intend to extrapolate the same in rabbits and other farm animals. In this method the transgene is delivered locally in the mammary gland hence devoid of hazard of germ line transmission of integrated transgene.
Apart from these we are developing methods for receptor mediated delivery of transgene through various nanoparticle based on Chitosan, Polyethylene Imine (PEI) tagged with transgene to transfect the mammary epithelial cells directly. We have standardized culture of primary mammary epithelial cells from mammary gland and milk of goat followed by invitro testing of these formulations to select the best transfecting agent for the invivo delivery of transgene.
Figure: A: Image showing Immuno Cyto Chemistry of Cultured Primary Mammary Epithelial Cells of Goat for Detecting CytoKeratine18 Expression (Marker for Mammary Luminal Epithelial Cells). B: Image showing expression of EGFP in primary Goat Mammary Epithelial cells transfected with EGFP expressing transgene construct integrated with Poly Ethelene Imine (PEI) Nanoparticle.
Using Ttransgenic Technology or Direct Transfection of Mammary Gland we are working to express cattle Follicle Stimulating Hormone (FSH) and Luteinizing Hormone (LH) in the Animal Bioreactor through Milk. We are also working for expressing human therapeutic protein Interferon-γ, Factor-8 and Tissue Plasminogen Activator using the same system.
Engineering Genome of Indian Livestock followed by Germ Cells/Spermatogonial Stem Cells (SSC) Transplantation to Enhance Productivity in Farm Animals:
Germ cell transplantation being an efficient tool for transgenesis are well studied in mice. Its application in farm animal is not well established though it has enormous potential to be used along with modern day genome editing tool CRISPR-CAS9 system, for editing genome of farm animals. We have developed method for evacuation of testis from endogenous germ cell population followed by transplantation of exogenous germ cells facilitating generation of sperm followed by progeny with donor derived genotype/phenotype. We are working towards exploring the possibilities of adopting this technology for the farm animals (Goat, Buffalo etc.). The work focuses on isolation and culture of germ cells, followed by engineering the genome of these cells using CRISPR-Cas9 system in-vitro, to introduce elite characteristic and transplanting them back into the evacuated testis.
Figure: A: Showing expression of DDX4, a germ cell marker in the testicular section of goat. B: showing colony of Spermatogonial Stem Cells (SSC) isolated from goat testis.
Using Multiplex CRISPR – Cas9 based genome editing we are Generating cells with knock-out of genomic region with undesired trait as well as cells with Site Specific Knock-in of DNA fragment with desired trait. Along with this, we are also adopting conventional shRNA based knock-down methods to silence the expression of gene in germ cells of farm animals.
Figure: Image showing Droplet Digital PCR analysis for detection of copy number in cultured primary epithelial cells of goat transfected with various multiplex CRISPR-Cas based construct to knock-out endogenous gene locus. Wildtype: denotes desired wildtype allele. CG_KO: denotes deletion of complete genomic region of the desired allele. EX_1+3_KO: denotes deletion of Exon1 and 3 of the desired allele. EX_1_KO: denotes deletion of Exon1 of the desired allele. EX_3_KO: denotes deletion of Exon3 of the desired allele. NTC: denotes no template control.
By this method we seek to increase productivity in farm animals as well as utilize the non-elite bulls to produce sperm with elite characteristic. Further we are also working to explore the alternative sources for generation of stem cells for several utility in farm animals by developing induced pluripotent stem cells (iPSCs) from somatic cells of farm animals.
Investigating Genetic Basis of Udder Gland Development:
The mammary gland is a dynamic organ that undergoes dramatic physiological adaptations during the life cycle specifically at the time of pregnancy to lactation. The mammary gland, and the genetic control of lactation (specifically milk production), evolved as a vital part of the mammalian development. Milk provides an essential source of nutrients to newborn mammals, as well as immune factors. Humans have long exploited the production of milk by domestic/farmed ruminants for the manufacture of dairy products, making milk an important part of human nutrition. The mammary gland displays a high level of developmental plasticity by undergoing repeated cycles of growth, differentiation, and regression, coordinated by the reproductive state. The ability to manipulate lactational output (specifically milk production) is an area of increasing interest. Knowledge of the biological pathways and mechanisms that govern mammary gland development and lactational yield is commercially important. We plan to undertake Next Generation Sequencing (NGS) of whole transcriptome from Mammary Gland of Indian Goat for detecting differentially expressed mRNA/Micro RNA (miRNA)/Long noncoding RNA (lncRNA) in different lactational time followed by Functional Genomics Study of such transcripts in transgenic mice models.
1. Goutam Ulgekar, Dilpreet Kaur, Venkateswaran Ganesan, Souvik Sen Sharma, Nirmalya Ganguli*, Subeer S Majumdar*. Anhydride chemistry based Hexanoylation of polyethylenimine increases transfection efficiency and expression of tagged DNA for therapeutic proteins in cultured cells. Biotechnol Bioeng, 2022 Nov;119(11):3275-3283. doi: 10.1002/bit.28196. Epub 2022 Aug 6. (*Co-Correspondence Author)
2. Anuradha Mishra, Nirmalya Ganguli, Subeer S Majumdar, and Deepak N Modi.Loss of HOXA10 causes endometrial hyperplasia progressing to endometrial cancer. Journal of Molecular Endocrinology. 2022. doi.org/10.1530/JME-22-0051
3. Abhishek Das, Satarupa Dutta, Dewanshu Sharma, Amit Pal, Nirmalya Ganguli*, Subeer S. Majumdar*.An easy method for developing fusion enabled SARS-CoV2 virus fusion mimic (SCFM), bypassing the need of Bio Safety Level (BSL) facility. Bioengineered, 2021, 12:1, 4407-4419, DOI: 10.1080/21655979.2021.1955509. (*Co-Correspondence Author)
4. Nirmalya Ganguli, Nilanjana Ganguli, Sunandini Chandra, Mayank Chaubey, Debi P. Sarkar and Subeer S. Majumdar. A Combinatorial Approach for Robust Transgene Delivery and Targeted Expression in Mammary Gland for Generating Biotherapeutics in milk, Bypassing Germline Gene Integration. Jouranl of Applied Microbiology and Biotechnology. 2018 Jul;102(14):6221-6234. doi: 10.1007/s00253-018-9094-2.
5. Nirmalya Ganguli, Neerja Wadhwa, Abul Usmani, Neetu Kunj, Nilanjana Ganguli, Rajesh Kumar Sarkar, Soma M Ghorai, Subeer S Majumdar. An efficient method for generating a germ cell depleted animal model for studies related to spermatogonial stem cell transplantation. Stem Cell Research and Therapy, 2016, 7: 142. doi:10.1186/s13287-016-0405-1
6. Abul Usmani*, Nirmalya Ganguli*, Subodh K Jain, Nilanjana Ganguli, Rajesh Kumar Sarkar, Mayank Choubey, Mansi Shukla, Hironmoy Sarkar and Subeer S Majumdar. Robust generation of transgenic mice by simple hypotonic solution mediated delivery of transgene in testicular germ cells. Molecular Therapy- Methods & Clinical Development, 2016, 3, 16076; doi:10.1038/mtm. 2016.76. (*Co 1st Author)
7. Nirmalya Ganguli, Nilanjana Ganguli, Abul Usmani and Subeer S. Majumdar. Isolation and functional characterization of buffalo (Bubalus bubalis) β-casein promoter for driving mammary epithelial cell-specific gene expression. Journal of Biotechnology, 2015, 198, 53-59.
8. Abul Usmani, Nirmalya Ganguli, Hironmoy Sarkar, Suveera Dhup, Suryaprakash R. Batta, Manoj Vimal, Nilanjana Ganguli, Sayon Basu, P. Nagarajan and Subeer S. Majumdar. A non-surgical approach for male germ cell mediated gene transmission through transgenesis. Scientific Reports, 2013, 3, Article number: 3430 doi: 10.1038/srep03430.
9. Richa Tiwari, Nirmalya Ganguli, Hunain Alam, Indrajit Sahu, Chella Krishna Vadivel, Shruti Sinha, Shweta Patel, Sayli Nitin Jamghare, Sanjay Bane, Rahul Thorat, Subeer S. Manjumdar, Milind M. Vaidya. Generation of a tissue‐specific transgenic model for K8 phosphomutants: A tool to investigate the role of K8 phosphorylation during skin carcinogenesis in vivo. Cell Biology International, 2021; doi.org/10.1002/cbin.11611.
10. Mansi Shukla, Nirmalya Ganguli, Souvik Sen Sharma, Subeer S Majumdar. Sertoli cell specific decline in NOR‐1 leads to germ cell apoptosis and reduced fertility. J Cell Biochem. 2018; 1–13. DOI: 10.1002/jcb.26698.
11. Sayon Basu, Satya Pal Arya, Abul Usmani, Bhola Shankar Pradhan, Rajesh Kumar Sarkar, Nirmalya Ganguli, Mansi Shukla, Kamal Mandal, Surendra Singh, Kanchan Sarda, Subeer S Majumdar. Defective Wnt3 expression by testicular Sertoli cells compromise male fertility. Cell and tissue research, 2018; 371 (2), 351-363.
1. Generating Transgenic Animal Models: Recent Technological Advancements.Neerja Wadhwa, Nirmalya Ganguli, Subeer S. Majumdar. In Book: Essentials of Laboratory Animal Science: Principles and Practices. Springer, Edited By, P. Nagarajan, Ramachandra Gudde, Ramesh Srinivasan. July 2021, DOI: 10.1007/978-981-16-0987-9_29
2. Animal Transgenesis in the era of Omics. In: Recent Advances in Communicable and non-communicable diseases. Edited by A. Datta and V. P. Sharma. Subeer S. Majumdar, Abul Usmani, Nirmalya Ganguli and Hironmoy Sarkar. (Capital Publishing Company, 2016, ISBN: 978-93-81891-31-5)
3. Transgenic animals: need of the country. Subeer Majumdar, Nirmalya Ganguli, Hironmoy Sarkar, Bhola Shankar Pradhan and Abul Usmani. INSA 100 lectures E book, Indian National Science Academy, New Delhi, India.
Title of the patent:
Hexanoyl modified high molecular weight polyethyleneimine for robust transfection in mammalian cells. Application No.: 202141033135
Current LAB Members:
Anandhi. R, (DBT-RA)
Ph. D. (Animal Biotechnology), Bharatidasan University, Tamilnadu.
Soma Behera (DBT-Senior Project Associate)
Ph. D. (Biochemistry), University of Hyderabad, Hyderabad.
Ph. D. Students:
Srimoyee Koner (JRF-UGC)
M.Sc. (Biotechnology), Presidency University, Kolkata.
Satarupa Dutta (DBT-Project Associate-II)
M.Sc. (Physiology), Calcutta University, Kolkata.
Amisha Bhattacharya (DST-Project Associate-I)
M.Sc. (Biotechnology), Amrita University, Kerala.
Harshada Thawari (DST-Project Associate-I)
M.Sc. (Animal Biology and Biotechnology), University of Hyderabad, Hyderabad.
DBT Project Associate-II
DBT Project Associate-II
DST Project Associate-I
DBT Project Associate-II
DBT Project Associate-II
1. Therapeutic protein production in milk of farm animals to increase their affordability.
2. Gene editing for generating tissue specific complete knock down/out of Myostatin gene for increased lean meat production in Indian goat (Capra hircus, Osmanabadi breed), Phase-I.
3. Feasibility of producing cattle gonadotropins in milk of rabbit by invivo gene transfection.
4. Differential transcriptome analysis in various stages of mammary gland development in goat.
5. An attempt to generate transgenic pig through testicular transgenesis or male germ cell transplantation to enhance productivity.
6. Targeted sorting of transgenic sperm for enhancing adaptability of testicular transgenesis.
7. Development of transgenic chicken as bioreactor for easy and cost effective production of human therapeutic proteins – tissue plasminogen activator (htPA) and erythropoietin (hERP).
1. Establishment of goat mammary epithelial/stem cell lines for the production of pharmaceutical interest proteins.
Dr. Nirmalya Ganguli,
Scientist – C
Gene and Protein Engineering Lab
National Institute of Animal Biotechnology
Survey No. 37, Opp. Journalist Colony
Extended Q City Road, Near Gowlidoddy
Telangana – 500032
Interested students can send email me for Ph. D. Position, Project Position and Dissertation with Statement of Purpose and Complete CV.