Means and methods for non-invasive diagnosis of chromosomal aneuploidy
The invention relates to a prenatal diagnostic method for the determination of a fetal chromosomal aneuploidy in a biological sample obtained from a pregnant woman, which method comprises enrichment and quantification of selected cell-free deoxyribonucleic acid sequences showing consensus nucleosome binding regions.
1. A method for determining a fetal chromosomal aneuploidy in a biological sample of a pregnant female individual, wherein the biological sample includes nucleic acid molecules, the method comprising:
(a) selecting for and isolating from a biological sample of a pregnant female individual one or more target sequences of DNA molecules present in the biological sample using sequence-specific selection of the target sequences, wherein said target sequences comprise DNA sequences including sequence regions located from 500 bp upstream to 1500 bp downstream of a transcription start site (TSS);
(b) amplifying said selected target sequences;
(c) sequencing said amplified selected target sequences, allotting each target sequence to a chromosome of the genome and identifying the unique allotted target sequences, wherein each identified unique allotted target sequence maps to a single chromosome;
(d) determining a first amount for each of one or more first chromosomes identified on the basis of said unique allotted target sequences originating from said one or more first chromosomes;
(e) determining a second amount for each of one or more second chromosomes identified on the basis of said unique allotted target sequences originating from said one or more second chromosomes; and
(f) determining based on the said first and second amount a fetal chromosomal aneuploidy of one or more of said first chromosomes.
2. The method of claim 1 , wherein step (f) further comprises:
(i) determining a parameter from said first amount relative to said second amount;
(ii) comparing the parameter to a corresponding cut off control value; and based on the comparison, determining whether or not there is a difference allowing for the prediction of a fetal chromosomal aneuploidy of one or more of said first chromosomes.
3. The method of claim 1 , wherein said biological sample is a maternal blood sample.
4. The method of claim 3 , wherein said blood sample is a sample of maternal blood plasma or maternal blood serum.
5. The method of claim 1 , wherein said biological sample is a urine sample or a saliva sample.
6. The method of claim 1 , wherein said one or more first chromosomes are selected from the group consisting of chromosome 21, chromosome 18, chromosome 13, chromosome X, and chromosome Y.
7. The method of claim 1 , wherein step (a) comprises contacting the biological sample with a nucleic acid probe specific for each target sequence.
8. The method of claim 7 , wherein the nucleic acid probe comprises RNA or DNA.
9. The method of claim 7 , wherein the nucleic acid probe is biotinylated.