Compositions and methods for multiplexed quantitative analysis of cell lineages

Abstract

Compositions and methods are provided for measuring population size for a plurality of clonal cell populations in the same individual, e.g., for measuring tumor size for a plurality of clonally independent tumors within the same individual. A subject method can include: (a) contacting an individual with a plurality of cell markers that are heritable and distinguishable from one another, to generate a plurality of distinguishable lineages of heritably marked cells; (b) after sufficient time has passed for the heritably marked cells to undergo at least one round of division, detecting and measuring quantities of at least two of the plurality of cell markers present in the contacted tissue, thereby generating a set of measured values; and (c) using the set of measured values to calculate the number of heritably marked cells that are present (e.g., for at least two of the distinguishable lineages of heritably marked cells).

1 - 54 . (canceled) 
     
     
         55 . A nucleic acid molecule comprising a multi-component barcode, wherein a first component of the barcode is unique to a portion of the nucleic acid molecule capable of causing a genetic perturbation of at least one gene in a recipient cell, and wherein a second component of the barcode is unique to the nucleic acid molecule. 
     
     
         56 . The nucleic acid molecule of  claim 55 , wherein the genetic perturbation of at least one gene in a recipient cell affects neoplastic cell formation. 
     
     
         57 . The nucleic acid molecule of  claim 55 , comprising one or more of: homology directed repair (HDR) DNA donor templates, nucleic acids encoding one or more oncogenes, nucleic acids encoding one or more wildtype proteins, nucleic acids encoding one or more mutant proteins, nucleic acids encoding one or more CRISPR/Cas guide RNAs, nucleic acids encoding one or more short hairpin RN As (shRNAs), or nucleic acids encoding one or more genome editing proteins. 
     
     
         58 . The nucleic acid molecule of  claim 57 , wherein the genome editing protein is selected from: a CRISPR/Cas RNA-guided protein, a CRISPR/Cas RNA-guided protein fused to a transcriptional activator or repressor polypeptide, a Cas9 protein, a Cas9 protein fused to a transcriptional activator or repressor polypeptide, a zinc finger nuclease (ZFN), a TALEN, a phage-derived integrase, a Cre protein, a Flp protein, and a meganuclease protein. 
     
     
         59 . The nucleic acid molecule of  claim 58 , wherein the genome editing protein comprises a CRISPR/Cas RNA-guided protein. 
     
     
         60 . The nucleic acid molecule of  claim 55 , wherein the at least one gene is a tumor suppressor gene. 
     
     
         61 . The nucleic acid molecule of  claim 55 , wherein the second component of the barcode is capable of distinguishing lineages of heritably marked cells. 
     
     
         62 . The nucleic acid molecule of  claim 61 , wherein the second component of the barcode comprises random nucleotides. 
     
     
         63 . The nucleic acid molecule of  claim 61 , wherein the second component of the barcode comprises defined nucleotides. 
     
     
         64 . The nucleic acid molecule of  claim 55 , further comprising a portion that encodes cre-recombinase. 
     
     
         65 . The nucleic acid molecule of  claim 55 , wherein the multi-component barcode comprises two components. 
     
     
         66 . The nucleic acid molecule of  claim 55 , wherein the multi-component barcode comprises three components. 
     
     
         67 . The nucleic acid molecule of  claim 55 , wherein the multi-component barcode comprises four components. 
     
     
         68 . The nucleic acid of  claim 55 , wherein the recipient cell is mammalian cell. 
     
     
         69 . A viral vector comprising the nucleic acid molecule of  claim 55 . 
     
     
         70 . The viral vector of  claim 69 , wherein the viral vector selected from the group consisting of a lentiviral vector, an adenoviral vector, an adeno-associated viral vector, a retroviral vector, a bocavirus vector, and a foamy virus vector. 
     
     
         71 . The viral vector of  claim 70 , wherein the vector is a plasmid. 
     
     
         72 . A genetically modified mouse comprising the nucleic acid molecule of  claim 55 . 
     
     
         73 . A genetically modified mouse comprising the viral vector of  claim 70 . 
     
     
         74 . A vertebrate tissue comprising the nucleic acid molecule of  claim 55 . 
     
     
         75 . A vertebrate tissue comprising the viral vector of  claim 70 .