# Establishing CRISPR cell screening for the deer tick Ixodes scapularis, a vector of Lyme and other diseases

> **NIH NIH R21** · HARVARD MEDICAL SCHOOL · 2022 · $263,825

## Abstract

Project Summary/Abstract
Prevention of tick-borne illnesses would be significantly aided by the availability of new strategies for
uncovering the functions of tick genes. Ixodes scapularis is an arthropod vector of the Lyme disease spirochete
Borrelia burgdorferi and other emerging human pathogens, including the rickettsial agent Anaplasma
phagocytophilum, the flavivirus Powassan, and the parasite Babesia microti. Unfortunately, genetic studies of
ticks are significantly hindered by practical barriers, including the approximately two-year life cycle of I.
scapularis. An alternative approach is to perform CRISPR screening in cultured cells, which in other systems is
an established robust and large-scale approach to uncovering new information about cellular activities and
pathways. We have successfully developed a method for genome-wide CRISPR screening in insect cells that
is extensible to ticks. In this R21 application, we will establish a genome-wide pooled CRISPR cell screening
platform for I. scapularis cultured cells as a scientific resource for the community. Tick cell culture has been
extensively used to investigate microbial interactions and can be used to predict the complex physiology of I.
scapularis without the challenges associated with the long-life cycle of ticks. Specifically, we will use our
combined knowledge of I. scapularis cultured cells and CRISPR cell screening to: (i) design single guide RNAs
(sgRNAs) for CRISPR modification based on the reference genome sequence and other genome sequences
of this species; (ii) identify appropriate U6 promoters for sgRNA expression in I. scapularis cells; (iii) modify
cells for screening, such as by making the competent for recombination mediated cassette exchange (RMCE)
and by introducing Cas9; and (iv) test and optimize the efficiency of CRISPR-based knockout strategies in
these cells. Concurrent with this work, we will develop cell-based assays appropriate for pooled-format screens
that interrogate tick immune signaling via the immune defense (IMD) and JAK/STAT pathways. Notably, the
type of large-scale and unbiased approach we propose will provide an important complement to reverse
genetic in vivo analyses. Establishment of CRISPR screening in I. scapularis cells will propel the field of tick
biology forward at a rapid pace, provide novel insights into relationships between arthropod vectors and the
microbes they host, foster collaborations between investigators with distinct expertise, and offer new
opportunities for mentoring the next generation of scientists.

## Key facts

- **NIH application ID:** 10428801
- **Project number:** 1R21AI168592-01
- **Recipient organization:** HARVARD MEDICAL SCHOOL
- **Principal Investigator:** STEPHANIE E MOHR
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $263,825
- **Award type:** 1
- **Project period:** 2022-08-18 → 2024-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10428801

## Citation

> US National Institutes of Health, RePORTER application 10428801, Establishing CRISPR cell screening for the deer tick Ixodes scapularis, a vector of Lyme and other diseases (1R21AI168592-01). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10428801. Licensed CC0.

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