PXD063746-1

PXD063746 is an original dataset announced via ProteomeXchange.

Title	Automation of a CRISPRi platform for enhanced isoprenol production in Pseudomonas putida [DBTL6]
Description	Recent advances in genome engineering technologies have significantly enhanced the ability to perturb microbial metabolic networks. Despite this progress, many bioproduction efforts struggle to parse complex metabolic datasets (e.g., -omics) within the constraints of the traditional design-build-test-learn (DBTL) cycle to efficiently optimize titer, rate, and yield. Here, we consider bioproduction of isoprenol, a precursor to a sustainable aviation fuel blendstock, in Pseudomonas putida. Initially, a combination of heuristics and genome scale metabolic modeling was used to identify 120 genes associated with isoprenol production for CRISPR interference. We then developed a largely automated conversion pipeline to design multiplexed guide RNA arrays, construct strains harboring these arrays, and then assess proteomics-validated isoprenol production in P. putida, which reduced the DBT turnaround time from months using conventional methods to two weeks. By leveraging an active learning model, the Automated Recommendation Tool (ART), we systematically and rapidly identified new experimental designs that improved titer. ART successfully downselected from an initial design space of 2.1 x 108 possible combinations to approximately 300 combinations. After 6 successive DBTL cycles this ultimately yielded several different gRNA combinations with up to 5-fold titer improvement. Our conversion pipeline not only showcases the potential of integrating laboratory automation with machine learning, but also marks a significant advancement toward autonomous strain design.
HostingRepository	PRIDE
AnnounceDate	2025-09-19
AnnouncementXML	Submission_2025-09-19_09:09:15.748.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Christopher Petzold
SpeciesList	scientific name: Pseudomonas putida KT2440; NCBI TaxID: 160488;
ModificationList	monohydroxylated residue; deamidated residue; iodoacetamide derivatized residue
Instrument	Orbitrap Exploris 480

Revision	Datetime	Status	ChangeLog Entry
0	2025-05-07 17:04:03	ID requested
⏵ 1	2025-09-19 09:09:16	announced

Dataset with its publication pending

submitter keyword: machine learning, SAF, isoprenol, JBEI,laboratory automation, CRISPRi

Christopher Petzold
contact affiliation	Staff Scientist Biological Systems & Engineering Division Lawrence Berkeley National Laboratory Berkeley CA 94720
contact email	cjpetzold@lbl.gov
lab head
Christopher Petzold
contact affiliation	Lawrence Berkeley National Laboratory
contact email	cjpetzold@lbl.gov
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD063746
     1. Label: PRIDE project
     2. Name: Automation of a CRISPRi platform for enhanced isoprenol production in Pseudomonas putida [DBTL6]