Simultaneous quantification of nucleic acids in diseased cells

Abstract

A process for assessing mitochondrial toxicity of a compound that includes contacting nucleic acids from a host with an amplification reaction mixture that contains at least two primers that provide detectable signals, wherein: a first primer provides a first detectable signal upon amplification of a host mitochondrial nucleic acid; a second primer provides a second detectable signal upon amplification of a host nuclear nucleic acid; and comparing the first and second detectable signals.

1 - 22 . (canceled)  
     
     
         23 . A process for assessing mitochondrial toxicity of a compound that includes: 
 contacting nucleic acids from a host with an amplification reaction mixture that contains at least two primers that provide detectable signals, wherein a first primer provides a first detectable signal upon amplification of a host mitochondrial nucleic acid; a second primer provides a second detectable signal upon amplification of a host nuclear nucleic acid;    and    comparing the first and second detectable signals.    
     
     
         24 . The process of  claim 23 , wherein the host mitochondrial nucleic acid is mitochondrial DNA.  
     
     
         25 . The process of  claim 23 , wherein the host mitochondrial nucleic acid is mitochondrial RNA.  
     
     
         26 . The process of  claim 23 , wherein the host mitochondrial nucleic acid is a non-coding sequence.  
     
     
         27 . The process of  claim 26 , wherein the non-coding sequence is a 5′-non-coding sequence.  
     
     
         28 . The process of  claim 26 , wherein the non-coding sequence is a 3′-non-coding sequence.  
     
     
         29 . The process of  claim 26 , wherein the non-coding sequence is an intron.  
     
     
         30 . (canceled)  
     
     
         31 . (canceled)  
     
     
         32 . The process of  claim 23 , wherein the host mitochondrial nucleic acid is a coding sequence.  
     
     
         33 . The process of  claim 23 , wherein the host nuclear nucleic acid is DNA.  
     
     
         34 . The process of  claim 23 , wherein the host nuclear nucleic acid is RNA.  
     
     
         35 . The process of  claim 23 , wherein the host nuclear nucleic acid is a nuclear non-coding sequence.  
     
     
         36 . The process of  claim 35 , wherein the nuclear non-coding sequence is a 5′-non-coding sequence.  
     
     
         37 . The process of  claim 35 , wherein the nuclear non-coding sequence is a 3′-non-coding sequence.  
     
     
         38 . The process of  claim 35 , wherein the nuclear non-coding sequence is an intron.  
     
     
         39 . The process of  claim 35 , wherein the nuclear non-coding sequence is from a gene part of which codes for β-actin.  
     
     
         40 . The process of  claim 35 , wherein the nuclear non-coding sequence is from a gene part of which codes for GAPDH.  
     
     
         41 . The process of  claim 23 , wherein the host nuclear nucleic acid is a coding sequence.  
     
     
         42 . The process of  claim 23 , wherein the second primer comprises SEQ ID No. 1.  
     
     
         43 . The process of  claim 23  further comprising wherein at least one detectable signal is provided upon interaction of at least two primers, wherein the detectable signal is caused by the hybridization of a third primer with a detectable agent to a primer that does not have a detectable agent which is hybridized to a host nucleic acid sequence.  
     
     
         44 . The process of  claim 43 , wherein the third primer comprises SEQ ID No. 2.  
     
     
         45 . The process of  claim 44 , wherein the reaction mixture further comprises a fourth primer.  
     
     
         46 . The process of  claim 45 , wherein the fourth primer which comprises SEQ ID No. 3.  
     
     
         47 . The process of  claim 46  wherein the reaction mixture further comprises a reporter and a quencher molecule.  
     
     
         48 . The process of  claim 47  wherein the reporter molecule is FAM and the quencher molecule is TAMRA.  
     
     
         49 . The process of  claim 43 , wherein the first primer comprises SEQ ID No. 19.  
     
     
         50 . The process of  claim 49 , wherein the reaction mixture further comprises a fifth primer.  
     
     
         51 . The process of  claim 50 , wherein the fifth primer comprises SEQ ID No. 20.  
     
     
         52 . The process of  claim 51 , wherein the reaction mixture further comprises a sixth primer.  
     
     
         53 . The process of  claim 52 , wherein the sixth primer comprises SEQ ID No. 21.  
     
     
         54 . The process of  claim 53  wherein the reaction mixture further comprises a reporter and a quencher molecule.  
     
     
         55 . The process of  claim 54  wherein the reporter molecule is TET and the quencher molecule is TAMRA.  
     
     
         56 . A kit for assessing mitochondrial toxicity of a compound, comprising a mixture of oligonucleotides comprising at least one first primer that provides a detectable signal on the occurrence of amplification of mitochondrial nucleic acid; and at least one second primer that provides a second detectable signal on the occurrence of amplification of host nuclear nucleic acid.  
     
     
         57 . A kit as in  claim 56  wherein the second primer comprises SEQ ID No. 1 and SEQ ID No. 2.  
     
     
         58 . A kit as in  claim 57  further comprising SEQ ID No. 3.  
     
     
         59 . A kit as in  claim 58  further comprising a fluorescent dye and a quenching dye.  
     
     
         60 . A kit as in  claim 56  wherein the first primer comprises SEQ ID No. 19.  
     
     
         61 . A kit as in  claim 60  further comprising a third primer that comprises SEQ ID No. 20.  
     
     
         62 . A kit as in  claim 61  further comprising a sixth primer that comprises SEQ ID No. 21.  
     
     
         63 . A kit as in  claim 62  further comprising a fluorescent dye and a quenching dye.