Massachusetts Institute of Technology

Harvard College

Shahar Alon

Edward Stuart Boyden

Fei Chen

Evan R Daugharthy

George M Church

Adam H Marblestone

Asmamaw T Wassie

Andrew C Payne

Paul Warren Tillberg

Jonathan Scheiman

Jason Daniel Buenrostro

Paul Reginato

Anubhav Sinha

Daniel Goodwin

The present invention provides methods for analyzing polynucleotides such as genomic DNA. In some embodiments, the disclosure provides a method for preparing and amplifying a genomic DNA library in situ in a fixed biological sample. The method comprises treating a fixed biological sample with an insertional enzyme complex to produce tagged fragments of genomic DNA. The method further comprises circularizing the tagged fragments of genomic DNA. The method further comprises amplifying the tagged fragments of genomic DNA.

In situ ATAC sequencing

The present invention relates to a method for amplifying at least one target RNA in a fixed and, optionally, expanded biological sample. In an embodiment of the invention, the method comprises incubating the fixed biological sample with a pair of polynucleotides complementary to non-overlapping and proximal sequences of a target RNA, wherein the polynucleotide pair hybridizes to the target RNA; ligating the polynucleotide pair using a ligase; and amplifying the ligation product. The invention further provides methods for detecting and optionally quantifying and/or sequencing the amplification product. As the method comprises hybridizing polynucleotide pairs to a target RNA in a fixed biological sample, the target RNA can be hybridized in situ.

Multiplexed signal amplified FISH via splinted ligation amplification and sequencing

The invention provides in situ nucleic acid sequencing to be conducted in biological specimens that have been physically expanded. The invention leverages the techniques for expansion microscopy (ExM) to provide new methods for in situ sequencing of nucleic acids as well as new methods for fluorescent in situ sequencing (FISSEQ) in a new process referred to herein as 'expansion sequencing' (ExSEQ).

In situ nucleic acid sequencing of expanded biological samples

The invention provides in situ nucleic acid sequencing to be conducted in biological specimens that have been physically expanded. The invention leverages the techniques for expansion microscopy (ExM) to provide new methods for in situ sequencing of nucleic acids in a process referred to herein as 'expansion sequencing' (ExSEQ).

Augmenting in situ nucleic acid sequencing of expanded biological samples with in vitro sequence information

The invention enables in situ genomic and transcriptomic assessment of nucleic acids to be conducted in biological specimens that have been physically expanded. The invention leverages the techniques for expansion microscopy (ExM) to provide new methods for in situ genomic and transcriptomic assessment of nucleic in a new process referred to herein as 'expansion fluorescent in situ hybridization' (ExFISH).

Nanoscale imaging of proteins and nucleic acids via expansion microscopy

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Cambridge, MA, United States of America

Shahar Alon