Non-invasive detection of fetal genetic traits

Abstract

Blood plasma of pregnant women contains fetal and (generally>90%) maternal circulatory extracellular DNA. Most of said fetal DNA contains ≦500 base pairs, said maternal DNA having a greater size. Separation of circulatory extracellular DNA of <500 base pairs results in separation of fetal from maternal DNA. A fraction of a blood plasma or serum sample of a pregnant woman containing, due to size separation (e.g. by chromatography, density gradient centrifugation or nanotechnological methods), extracellular DNA substantially comprising ≦500 base pairs is useful for non-invasive detection of fetal genetic traits (including the fetal RhD gene in pregnancies at risk for HDN; fetal Y chromosome-specific sequences in pregnancies at risk for X chromosome-linked disorders; chromosomal aberrations; hereditary Mendelian genetic disorders and corresponding genetic markers; and traits decisive for paternity determination) by e.g. PCR, ligand chain reaction or probe hybridization techniques, or nucleic acid arrays.

1 - 21 . (canceled) 
     
     
         22 . Isolated deoxyribonucleic acid (DNA) from blood plasma of a pregnant female and enriched for fetal DNA, which isolated DNA consists of:
 DNA fragments comprising fetal DNA,   which DNA fragments are not in association with a gel,   which fetal DNA comprises at least one allele different than a maternal allele, and   which isolated DNA is prepared by a process comprising:   (a) extracting circulating extracellular DNA from the plasma from the pregnant female, thereby providing extracted DNA; and   (b) enriching the extracted DNA for DNA fragments of less than 500 base pairs, thereby providing enriched DNA;   whereby the enriched DNA is enriched for fetal DNA.   
     
     
         23 . The isolated DNA of  claim 22 , wherein the extracted DNA is enriched for DNA fragments of less than 300 base pairs in (b). 
     
     
         24 . The isolated DNA of  claim 22 , wherein the fetal DNA is from a fetus having a chromosome aneuploidy. 
     
     
         25 . The isolated DNA of  claim 22 , wherein the fetal DNA comprises a paternally inherited polymorphism not present in a maternally inherited sequence and the paternally inherited polymorphism is of a length different than in the maternally inherited sequence. 
     
     
         26 . The isolated DNA of  claim 22 , wherein the DNA fragments are in solution. 
     
     
         27 . The isolated DNA of  claim 22 , wherein enriching the extracted DNA in (b) comprises electrophoresis. 
     
     
         28 . The isolated DNA of  claim 22 , wherein enriching the extracted DNA in (b) comprises chromatography. 
     
     
         29 . Isolated deoxyribonucleic acid (DNA) from blood plasma of a pregnant female and enriched for fetal DNA, which isolated DNA consists of:
 DNA fragments comprising fetal DNA,   which DNA fragments are not in association with a gel,   which fetal DNA comprises at least one allele different than a maternal allele, and   which isolated DNA is prepared by a process comprising:   (a) extracting circulating extracellular DNA from the plasma from the pregnant female, thereby providing extracted DNA; and   (b) selecting for DNA fragments of less than 500 base pairs from the extracted DNA, thereby providing selected DNA;   whereby the selected DNA is enriched for fetal DNA.   
     
     
         30 . The isolated DNA of  claim 29 , wherein DNA fragments of less than 300 base pairs are selected for in (b). 
     
     
         31 . The isolated DNA of  claim 29 , wherein the fetal DNA is from a fetus having a chromosome aneuploidy. 
     
     
         32 . The isolated DNA of  claim 29 , wherein the fetal DNA comprises a paternally inherited polymorphism not present in a maternally inherited sequence and the paternally inherited polymorphism is of a length different than in the maternally inherited sequence. 
     
     
         33 . The isolated DNA of  claim 29 , wherein the DNA fragments are in solution. 
     
     
         34 . The isolated DNA of  claim 29 , wherein selecting for DNA fragments in (b) comprises electrophoresis. 
     
     
         35 . The isolated DNA of  claim 29 , wherein selecting for DNA fragments in (b) comprises chromatography. 
     
     
         36 . Isolated deoxyribonucleic acid (DNA) from blood plasma of a pregnant female and enriched for fetal DNA, which isolated DNA consists of:
 DNA fragments comprising fetal DNA,   which DNA fragments are not in association with a gel,   which fetal DNA comprises at least one allele different than a maternal allele, and   which isolated DNA is prepared by a process comprising:   (a) extracting circulating extracellular DNA from the plasma from the pregnant female, thereby providing extracted DNA; and   (b) size discriminating the extracted DNA for DNA fragments of less than 500 base pairs, thereby providing size discriminated DNA;   whereby the size discriminated DNA is enriched for fetal DNA.   
     
     
         37 . The isolated DNA of  claim 36 , wherein DNA fragments of less than 300 base pairs are size discriminated in (b). 
     
     
         38 . The isolated DNA of  claim 36 , wherein the fetal DNA is from a fetus having a chromosome aneuploidy. 
     
     
         39 . The isolated DNA of  claim 36 , wherein the fetal DNA comprises a paternally inherited polymorphism not present in a maternally inherited sequence and the paternally inherited polymorphism is of a length different than in the maternally inherited sequence. 
     
     
         40 . The isolated DNA of  claim 36 , wherein the DNA fragments are in solution. 
     
     
         41 . The isolated DNA of  claim 36 , wherein the size discriminating in (b) comprises electrophoresis. 
     
     
         42 . The isolated DNA of  claim 36 , wherein the size discriminating in (b) comprises chromatography. 
     
     
         43 . Isolated deoxyribonucleic acid (DNA) from blood plasma of a pregnant female and enriched for fetal DNA, which isolated DNA consists of:
 DNA fragments less than 500 base pairs in length,   which DNA fragments are not in association with a gel,   a portion of which DNA fragments comprise fetal DNA and a portion of which DNA fragments comprise maternal DNA,   which fetal DNA comprises at least one allele different than a maternal allele in the maternal DNA, and   which fetal DNA comprises Y-chromosome specific sequences.   
     
     
         44 . The isolated DNA of  claim 43 , wherein the Y-chromosome specific sequences comprise an SRY sequence. 
     
     
         45 . The isolated DNA of  claim 44 , wherein the DNA fragments are less than 300 base pairs in length. 
     
     
         46 . The isolated DNA of  claim 44 , wherein the DNA fragments are in solution.