Pablo Mantilla Puccetti

Currently exploring the novel phage Pasto, specifically its MarR-like regulatory repressor that allows for the integration of Pasto’s genome into Agrobacterium sp. Research has consisted of inducing directed evolution, with techniques like the Appleman’s method, in phage Pasto to create virulent mutants, sequencing these mutants, and analyzing these sequences to derive what components of the MarR-like regulatory system are necessary for facilitating integration. Analysis of these mutants and wildtype sequences has led to multiple sequence alignments with tcoffee, homology searches with BLAST, quaternary structure analysis with Alphafold, and bacterial chromosome analysis with PHASTEST.

A Research Experience for Undergraduates (REU) program where I had the opportunity to investigate the impact of aging on the mammary gland's response to pregnancy under Dr. Camila dos Santos. I extracted mice's mammary glands and suspended them into organoids for hormonal treatment. I independently analyzed the resulting scRNA-seq data in R. Applying the Seurat and Cellchat pipelines in R, I clustered cells into distinct cell types based on their gene-expression similarity, allowing me to compare clusters via differential pathway enrichment and gene expression analysis to understand the differential effects of pregnancy hormones. I presented my project orally many times at lab meetings, preliminary talks, and the Cold Spring Harbor Laboratory Undergraduate Research Symposium. While assisting with graduate student’s projects, I familiarized myself with immunofluorescence staining and visualization, western blot, and cryostat tissue sectioning. In addition, the program offered many bioinformatics/computational biology courses where I was able to familiarize myself with some of the many biological applications of machine learning.

Here, I explored the protection provided by Spiroplasma poulsonii against the wasps Ganaspis sp. and Leptopilina sp. affinity clavipes when attempting to parasitize Drosophila melanogaster. Data was collected via protection experiments, which consisted of isolating a fixed number of fly pupae and wasps in individual “environments”, with the goal of quantifying wasp susceptibility to S.poulsonii. My main responsibilities here included planning and executing protection experiments, verifying fly infection status via PCR amplification/Gel electrophoresis, and teaching lab protocols to incoming members. Additionally, I did bioinformatics work on refining the shotgun sequencing of Poeciliopsis monacha and P.jackshultzi alleles in P.monacha-jackshultzi hybrids on HPRC Maroon Galaxy. Lab meetings and journal clubs were also part of the weekly schedule, which I would lead occasionally. My undergraduate research thesis highlighted the increased susceptibility to S.poulsonii of Ganaspis sp. when compared to Leptopilina sp. and the unexpectedly low oviposition frequency of Ganaspis sp.