Method of nuclear dna and mitochondrial dna analysis

Abstract

An amount of mitochondrial DNA molecules relative to an amount of nuclear DNA molecules is determined in a biological sample, and the relative amount is used for various purposes, e.g., screening, detection, prognostication or monitoring of various physiological and pathological conditions. As examples, an amount of mitochondrial DNA can be used to estimate a concentration of DNA of a tissue type, such as a fetal DNA concentration, tumor DNA concentration, or a concentration of DNA in the biological sample derived from a non-hematopoietic tissue source. Sequencing techniques can be used to determine a mitochondrial DNA concentration in a sample for an accurate detection of a level of cancer. A level of an auto-immune disease is also determined using a relative amount of mitochondrial DNA molecules compared nuclear DNA molecules.

What is claimed is: 
     
         1 . A method of analyzing a biological sample of an organism, the biological sample including cell-free DNA, the cell-free DNA including mitochondrial DNA and nuclear DNA, the method comprising:
 receiving, at a computer system, sequence information of a plurality of DNA molecules in the biological sample;   for each DNA molecule of the plurality of DNA molecules:
 determining, by the computer system, a location of the DNA molecule in a reference nuclear genome or a reference mitochondrial genome using the sequence information for the DNA molecule; 
 identifying, by the computer system, whether the DNA molecule is a nuclear DNA molecule or a mitochondrial DNA molecule based on the location; 
   measuring, by the computer system, a normalized amount of the plurality of DNA molecules that are mitochondrial DNA molecules, the normalized amount being relative to a second amount of the plurality of DNA molecules including DNA molecules that are identified as nuclear DNA molecules;   comparing the normalized amount to a reference value; and   determining a classification of a level an auto-immune disease in the organism based on the comparison.   
     
     
         2 . The method of  claim 1 , wherein determining the classification includes determining whether the auto-immune disease is active or inactive. 
     
     
         3 . The method of  claim 1 , wherein the normalized amount is compared to a plurality of reference values to determine the classification of the level of the auto-immune disease. 
     
     
         4 . The method of  claim 3 , wherein the plurality of reference values are part of a calibration function that provides a severity of the auto-immune disease. 
     
     
         5 . The method of  claim 4 , wherein the auto-immune disease is systemic lupus erythematosus (SLE), and wherein the severity is measured by a systemic lupus erythematosus disease activity index (SLEDAI). 
     
     
         6 . The method of  claim 4 , wherein the auto-immune disease is systemic lupus erythematosus (SLE), the method further comprising:
 measuring an anti-ds DNA antibody level in the biological sample, wherein the classification of the level of the auto-immune disease in the organism is further based the anti-ds DNA antibody level.   
     
     
         7 . The method of  claim 6 , further comprising:
 selecting the reference value based on the anti-ds DNA antibody level.   
     
     
         8 . The method of  claim 1 , further comprising performing, by the computer system:
 determining a first amount of the plurality of DNA molecules that are mitochondrial DNA molecules;   determining the second amount of the plurality of DNA molecules by counting the nuclear DNA molecules; and   computing a ratio of the first amount and the second amount, wherein the normalized amount of the plurality of DNA molecules that are identified as mitochondrial DNA molecules is determined using the ratio.   
     
     
         9 . A method of analyzing a biological sample of an organism, the biological sample including cell-free DNA, the cell-free DNA including mitochondrial DNA and nuclear DNA, the method comprising:
 receiving, at a computer system, sequence information of a plurality of DNA molecules in the biological sample;   for each DNA molecule of the plurality of DNA molecules:
 determining, by the computer system, a location of the DNA molecule in a reference mitochondrial genome using the sequence information for the DNA molecule; and 
 measuring, by the computer system, a size of the DNA molecule using the determined location of the DNA molecule; 
   identifying, by the computer system, a group of DNA molecules as being mitochondrial DNA molecules based on the determined locations;   calculating, by the computer system, a first statistical value of a size distribution of the group of mitochondrial DNA molecules;   comparing the first statistical value to a reference value; and   determining a classification of a level an auto-immune disease in the organism based on the comparison.   
     
     
         10 . The method of  claim 9 , wherein determining the classification includes determining whether the auto-immune disease is active or inactive based on the comparison. 
     
     
         11 . The method of  claim 9 , wherein the first statistical value is compared to a plurality of reference values to determine the classification of the level of the auto-immune disease. 
     
     
         12 . The method of  claim 11 , wherein the auto-immune disease is determined to be active when the first statistical value is greater than a first reference value, and wherein the auto-immune disease is determined to be inactive when the first statistical value is less than the first reference value and greater than a second reference value. 
     
     
         13 . The method of  claim 12 , wherein the auto-immune disease is determined to not exist when the first statistical value is less than the second reference value. 
     
     
         14 . The method of  claim 10 , wherein the first statistical value includes at least one of: an average, a median, or a mode of the size distribution of the group of mitochondrial DNA molecules. 
     
     
         15 . The method of  claim 9 , wherein the auto-immune disease is systemic lupus erythematosus (SLE).