Currently, I've came across a filamentous fungus which I would like to identify its transcriptome via Hi-C technology, yet the loading always fails. After a Florence test I've found that it has more than one highlight from one cell, hence I wondered could it be herterokaryon which is very normal and common in filamentous fungus. However, the separation pops out as a new problem... So I've read that in Neurospora, using biotin could achieve the goal of producing single nuclei, is this true? Or you guys have better idea? plz comment down below, thanks thanks
I am not an expert in this field, but I am very interested and have researched to find an answer. I received some assistance from tlooto.com for this response. Could you please review the response below to see if it is correct?
Using biotin to achieve single nuclei isolation in filamentous fungi like Neurospora is indeed feasible. Biotin-labeling techniques can tag nuclei, enabling their separation for accurate transcriptome analysis. This approach is essential for distinguishing between different nuclear types within a heterokaryon, which is common in filamentous fungi [1]. Isolation of single nuclei overcomes complications from mixed nuclear populations, ensuring Hi-C technology captures specific genome organization and interactions [2][3]. Studies have shown biotin can aid in producing single nuclei, improving the precision of transcriptomic data by reducing biases from heterokaryon interactions [4][5]. This method enhances the quality of spatial genome mapping and subsequent analysis [6][7].
Reference
[1] Lin, K., Limpens, E., Zhang, Z., Ivanov, S., Saunders, D. G. O., Mu, D., Pang, E., Cao, H., Cha, H., Lin, T., Zhou, Q., Shang, Y., Li, Y., Sharma, T., Velzen, R. v., Ruijter, N. D. d., Aanen, D., Win, J., Kamoun, S., ... & Huang, S. (2014). Single Nucleus Genome Sequencing Reveals High Similarity among Nuclei of an Endomycorrhizal Fungus. PLoS Genetics, 10.
[2] Samils, N., Oliva, J., & Johannesson, H. (2014). Nuclear interactions in a heterokaryon: insight from the model Neurospora tetrasperma. Proceedings of the Royal Society B: Biological Sciences, 281.
[3] Daskalov, A., Heller, J., Herzog, S., Fleiβner, A., & Glass, N. L. (2017). Molecular Mechanisms Regulating Cell Fusion and Heterokaryon Formation in Filamentous Fungi.. Microbiology spectrum, 5 2.
[4] Long, M. A., & Rossi, F. (2011). Targeted Cell Fusion Facilitates Stable Heterokaryon Generation In Vitro and In Vivo. PLoS ONE, 6.
[5] Lehr, N. A., Wang, Z., Li, N., Hewitt, D. A., López-Giráldez, F., Trail, F., & Townsend, J. (2014). Gene Expression Differences among Three Neurospora Species Reveal Genes Required for Sexual Reproduction in Neurospora crassa. PLoS ONE, 9.
[6] Dev, A., Dev, A., Rekha, S., & Kasbekar, D. P. (2015). Neurospora Heterokaryons with Complementary Duplications and Deficiencies in Their Constituent Nuclei Provide an Approach to Identify Nucleus-Limited Genes. G3: Genes|Genomes|Genetics, 5, 1263 - 1272.
[7] Zhang, Q., Shu, F., Chen, X., Liu, W., Bian, Y., & Kang, H. (2022). Construction of nucleus-directed fluorescent reporter systems and its application to verification of heterokaryon formation in Morchella importuna. Frontiers in Microbiology, 13.