WO2014101024A1 - Méthode de détermination sexuelle d'embryons de jumeaux, système et support lisible par ordinateur connexes - Google Patents

Méthode de détermination sexuelle d'embryons de jumeaux, système et support lisible par ordinateur connexes Download PDF

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WO2014101024A1
WO2014101024A1 PCT/CN2012/087582 CN2012087582W WO2014101024A1 WO 2014101024 A1 WO2014101024 A1 WO 2014101024A1 CN 2012087582 W CN2012087582 W CN 2012087582W WO 2014101024 A1 WO2014101024 A1 WO 2014101024A1
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sequencing data
twins
chromosome
fetuses
determined
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PCT/CN2012/087582
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English (en)
Chinese (zh)
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郑晶
张春雷
谢伟伟
蒋浩君
郭靖
杨焕明
张秀清
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深圳华大基因医学有限公司
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Priority to PCT/CN2012/087582 priority Critical patent/WO2014101024A1/fr
Publication of WO2014101024A1 publication Critical patent/WO2014101024A1/fr

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6879—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for sex determination

Definitions

  • This invention relates to the field of biomedicine and, in particular, to the field of genetic variation testing, and more particularly to methods, systems and computer readable shields for determining the sex of a fetus in a fetus. Background technique
  • Twins can be divided into identical twins and fraternal twins.
  • the development of two embryos from one fertilized egg is called an identical twin.
  • the twins developed from a single fertilized egg carry the same genetic background, and their gender, appearance, etc. are identical. If one of the fetuses has a genetic disease, the other fetus must be the same.
  • single-oval twins may be monochorionic mono-amnion, mono-chorionic double amnion, and double-chorioamnion amnion depending on the time of separation.
  • a fraternal twin is a discharge of two egg cells at a time, both of which develop into two embryos after fertilization.
  • fraternal twins randomly inherit the parent's genome according to Mendelian inheritance, and the types of genes carried are not identical. Therefore, they may have different genders and appearances, and the carrying status of genetic diseases may also be different.
  • the fraternal twins may also be double chorionic double amnion or monochorionic double amnion.
  • the detection techniques commonly used in prenatal and postnatal testing for twins are divided into non-invasive prenatal techniques and invasive prenatal techniques.
  • non-invasive prenatal technical damage is small, no trauma sampling is needed, but the current detection methods have disadvantages: 1) The risk factor of Down's child is calculated by checking the concentration of serum metabolites, the age of pregnant women, and the gestational age of blood stasis. However, the false positive rate and false negative rate of this Down's screening method are both high; 2) Ultrasound examination depends on the equipment and doctor's personal experience, can not form a unified standard, can not judge the developmental abnormalities of unclear or controversial ultrasound indicators .
  • the prenatal antenatal detection technology collects fetal samples with high accuracy, but it is easy to cause abortion, amniocentesis and other problems, which brings certain abortion risks to pregnant women and fetuses. It also has time limitations and invasive sampling. In a specific time zone. Moreover, the current methods are technically demanding and require lengthy experimental procedures and high costs.
  • the invention proposes a method of determining the sex of a fetus in a twin.
  • UR% determining the sex of the fetus in the twins, wherein at least one of the twins is determined to be a male fetus when at least one of the following conditions is satisfied, when none of the following conditions are satisfied.
  • Y-UR% is greater than the first threshold; and -1111% (-1; 11% is greater than the second threshold.
  • the invention proposes a computer readable medium.
  • the computer readable medium stores instructions adapted to be executed by the processor to determine the sex of the fetus in the twins by: obtaining a plurality of sequencing data of blood DNA of the twin pregnant women; Aligning the sequencing data with a human reference genomic sequence to obtain a unique aligned sequencing data set consisting of a plurality of unique aligned sequencing data; determining unique unique alignment sequencing data contained in the unique alignment sequencing data set a number M; determining the number N y of unique alignment sequencing data derived from the Y chromosome in the unique alignment sequencing data set, and at least one of the number N x of unique alignment sequencing data derived from the X chromosome; determining the chromosome
  • the sequencing data content of Y-UR% and the X-UR% sequencing data content of at least one of X-UR%, wherein, according to the formula Y-UR% N y /M, the Y-
  • X-UR% N X / M, to determine the content of the sequencing data of the X chromosome -UR%; and determining the sex of a fetus in twins, wherein, when the following conditions to When one of determining at least one of the twin male fetuses, when the following conditions are not satisfied, determining whether the twins are female fetuses: Y-UR 0 /. Greater than the first threshold; and ⁇ -13 ⁇ 4% «-13 ⁇ 4% is greater than the second threshold.
  • the computer-readable shield it is possible to effectively determine whether a male fetus is present in the twins, and then according to the fetal DNA concentration in the maternal plasma, the two fetus sexes can be effectively inferred.
  • the invention proposes a system for determining the sex of a fetus in a twin.
  • the system comprises: a sequencing device for sequencing blood DNA of a twin pregnant woman to obtain a plurality of sequencing data; a comparison device, the comparison device and the sequencing device Connected, the sequencing data is aligned with the reference genomic sequence of the organism to obtain a unique aligned sequencing data set consisting of a plurality of unique aligned sequencing data; an analysis device, the analysis device and the ratio Connected to the device and used to determine The fetal sex in the twins, wherein the analyzing device further comprises: a first calculating module, wherein the first calculating module is configured to determine a number M of unique aligned sequencing data included in the unique aligned sequencing data set; a calculation module, the second calculation module is configured to determine a number ⁇ N y of the unique alignment sequencing data derived from the ⁇ chromosome in the unique alignment sequencing data set, and
  • the invention also proposes a system for determining the sex of a fetus in a twin.
  • the system comprises: a sequencing device for sequencing blood DNA of the twin pregnant women to obtain a plurality of sequencing data; and the computer readable medium of the present invention as described above , wherein the computer readable shield stores instructions for determining the sex of the fetus in the twins.
  • the total number of sequencing data located on a chromosome is proportional to the length of the chromosome and the content of the chromosome in the biological sample, the system is used to determine the blood DNA of the pregnant woman of the twins.
  • the number of unique alignments of the X and Y chromosomes in the sample and the amount of sequencing data can effectively determine whether there are male fetuses in the twins, and based on the estimated fetal DNA concentration in the maternal plasma, the two fetus genders can be effectively inferred.
  • FIG. 1 shows a schematic flow chart of a method for determining the sex of a fetus in a twin, in accordance with one embodiment of the present invention
  • FIG. 2 shows a schematic structural view of a system for determining the sex of a fetus in a twin according to an embodiment of the present invention
  • Figure 3 shows the chromosome number of sample S 1 obtained by the method of the present invention, in accordance with one embodiment of the present invention.
  • Figure 4 shows the chromosomal digital karyotype results of sample S2 obtained using the method of the present invention, in accordance with one embodiment of the present invention. Detailed description of the invention
  • first and second are used for descriptive purposes only, and are not to be construed as indicating or implying a relative importance or implicitly indicating the number of technical features indicated.
  • the features of the first and second features may include one or more of the features, either explicitly or implicitly. Further, in the description of the present invention, unless otherwise stated, the meaning of "multiple" is two or More than two.
  • the invention proposes a method for determining the presence or absence of a male fetus in a twin.
  • the method can include:
  • the blood DNA of the twin pregnant women who need to be tested is first sequenced to obtain a plurality of sequencing data.
  • the method of extracting blood DNA of a pregnant woman of a twin is not particularly limited.
  • the blood DNA of the twin pregnant women can be extracted by at least one selected from the group consisting of magnetic bead method and column chromatography, preferably magnetic bead method.
  • the presence or absence of a male fetus in the twins can be effectively determined by the method of the present invention.
  • the principle of magnetic extraction of DNA by magnetic beads is as described above, and will not be described herein.
  • sequencing device that can be employed according to an embodiment of the present invention is not particularly limited.
  • sequencing is performed by at least one selected from the group consisting of Roche/454 GS Junior. Illumina/MiSeq and Life Tecnologies/Ion Torrent PGM, taking into account the advantages of instrument portability and high throughput performance. Thereby, it is possible to further improve the efficiency of determining whether or not a male fetus is present in the twins by utilizing the characteristics of high-throughput and deep sequencing of these sequencing devices.
  • the sequencing type can be single-end (one-way) sequencing or pair-end (two-way) sequencing. According to some embodiments of the invention, the sequenced data is 36 to 100 bp in length.
  • the sequencing platform employed is Ilhmiina/Solexa and the sequencing type is SE36 sequencing.
  • the efficiency of the subsequent analysis can be further improved.
  • the sequencing amount is a 5M sequencing short fragment.
  • the method for constructing the sequencing library can include: First, the DNA fragment is blunt-ended and the base A is added at the end, and the linker is ligated to obtain a DNA fragment having a linker;
  • the DNA having the adaptor is amplified to obtain an amplification product, that is, a sequencing library.
  • a tag sequence Index can be introduced into the sequencing library during the process of constructing the sequencing library, for example, an index can be introduced in the linker, or a tag sequence Index can be introduced during the amplification process.
  • simultaneous sampling of multiple test samples can be achieved by using different tag sequences for different samples.
  • the obtained sequencing data is aligned with a human reference genome sequence to obtain a unique alignment sequencing data set composed of a plurality of unique alignment sequencing data; and then, the unique alignment sequencing data is determined.
  • the number of unique aligned sequencing data contained in the set is M; then, the number N x and N y of unique aligned sequencing data derived from the X and Y chromosomes, respectively, in the unique aligned sequencing data set are determined.
  • the human reference genomic sequence used is a reference sequence obtained by masking the repeat sequence of the human genome sequence, such as the NCBI database.
  • the latest version of the Human Genome Reference Sequence is the human genome reference sequence of version 36 (hgl8; NCBI Build 36) in the NCBI database.
  • sequence alignment can be performed by any sequence alignment program, such as Short Oligonucleotide Analysis Package (SOAP) and BWA alignment (available by those skilled in the art) At least one of Burrows-Wheeler Aligner is performed to align the sequencing data with a reference genomic sequence to obtain the position of the sequencing data on the reference genome. Sequence alignment can be performed using default parameters provided by the program, or can be selected by those skilled in the art as needed.
  • the comparison software employed is SOAP aligner/soap2
  • the term "unique alignment sequencing data” refers to a sequence that has only a unique position on a reference genomic sequence when the sequencing data is aligned with a reference genomic sequence, as Unique reads.
  • Unique reads In an embodiment of the invention, in order to avoid interference from repetitive sequences, it is necessary to remove those DNA sequences that are located in tandem repeats and transposition repeat positions in the human genome reference sequence, and only those DNA sequences that can be mapped to unique positions in the genome are counted. That is, the only alignment of the sequencing data.
  • the unique alignment sequencing data maps the DNA sequences from the DNA of twin pregnant women to the specific chromosomes after being interrupted and sequenced.
  • the twins of the twin pregnant women can be determined based on the values of ⁇ -111% «-1; 1% and Y-UR%. Whether there is a male fetus. Specifically, when at least one of the following conditions is satisfied, it can be determined that at least one of the twins is a male fetus, and when none of the following conditions are satisfied, it can be determined that the twins are female fetuses: ⁇ -1!1% is greater than the first threshold ; and Y-UR%/X-UR% are greater than the second threshold.
  • the first threshold value and the second threshold value are determined by the following method: 100 samples of maternal fetal plasma samples are taken, and Y-UR is calculated separately. Y-UR%/X-U %, and draw the Y-UR of each sample separately. /. And the normal distribution curve of Y-UR%/X-UR%, according to the central limit theorem, the Y-UR% and Y-UR%/X-UR% of the 100 samples are in accordance with the normal distribution, taking Y-UR The right interval point of the °/W 99% confidence interval is taken as the first threshold, taking Y-UR%/X-UR°/. The right interval point of the 99% confidence interval is used as the second threshold.
  • the first threshold is determined to be 0.0004, and the second threshold is 0.0075.
  • Y-UR% is greater than 0.0004 or Y-UR%/X-UR% is greater than 0.0075, it can be determined that a male tire is present in the twin.
  • the method when it is determined that at least one of the twins is a male fetus, the method further includes: first, estimating the fetal DNA concentration based on the obtained N x and N Y respectively, so as to obtain the first fetus respectively L DNA M%chrX, and second fetal DNA concentration %chrY.
  • fetal DNA concentration estimation can be performed by the following steps: selecting normal male and normal female plasma sample sets as controls, respectively calculating ⁇ -111% and -UR% of each plasma sample; respectively calculating each male plasma Sample Y-UR 0 /.
  • the arithmetic mean of X-UR% was calculated for each female plasma sample as Y-UR°/ ⁇ n X-UR 0 /.
  • the arithmetic mean value denoted as Y F and X F ; then, the first fetal DNA M%chrX estimated using the X chromosome is calculated according to the formula (X-UR%-X F )/(X M -X F ), according to The formula (Y-UR%-Y F )/(Y M -Y F ) calculates the second fetal DNA concentration 3 ⁇ 4%chrY estimated using the Y chromosome, where -111% of the formula is the X of the plasma sample of the pregnant woman to be tested
  • the -UR% value, Y-UR% is the Y-UR% value of the plasma sample of the pregnant woman to be tested.
  • the significant difference p-values of the % ⁇ 13 ⁇ 4 and %(;1« ⁇ are determined. Further, the sex of the two fetuses in the twins can be determined based on the p-value. Specifically, according to an embodiment of the present invention, P-value>0.1, it can be determined that the fetus in the twins is a male fetus. If p-value ⁇ 0.1, and /chrX is greater than %chrY, it can be determined that there is one male fetus and one female fetus in the twin.
  • the concentration of DNA on the X and Y chromosomes contributed by the two fetuses will be equal (because the two fetuses provide two X chromosomes and two Y chromosomes); if the two fetuses are a male fetus and a female
  • the concentration of DNA on the X and Y chromosomes contributed by the two fetuses is that the DNA concentration on the X chromosome is higher than the DNA concentration on the Y chromosome (because of this When the two fetuses provide three X chromosomes and one Y chromosome).
  • the sex of the twin fetus can also be inferred by using the fetal DNA concentration estimated by the variant chromosome. Specifically, if the estimated fetal DNA concentration using the variant chromosome is about using the ⁇ chromosome Estimating the 1/2 of the fetal DNA concentration obtained, it is concluded that the two fetuses are two male fetuses.
  • the gender of the fetus is estimated by using the estimated fetal DNA concentration of the mutant chromosome, which is premised on the assumption that only one fetus is abnormal (because the probability of abnormality of both fetuses is small), and further, The fetal concentration estimated using abnormal chromosomes is only the DNA concentration provided by one of the fetuses. If both fetuses are male, then the fetal concentration estimated using the sputum chromosome is the DNA concentration provided by the two fetuses, theoretically twice the fetal concentration estimated using the mutated chromosome. Thus, when the fetal DNA concentration estimated using the mutant chromosome is about 1/2 of the fetal DNA concentration estimated using the sputum chromosome, it can be inferred that both fetuses are male fetuses.
  • the tag sequence Index can be introduced into the sequencing library during the process of constructing the sequencing library, thereby implementing different tag sequences for different samples.
  • multiple test samples are sequenced, and the sequencing data of each sample is effectively distinguished by the label, and then the values of M, N x and N y of each sample can be respectively obtained, so that whether there are male tires in the subsequent twins respectively Judgment.
  • the method of determining whether a male fetus is present in a twin may further comprise: statistically comparing the number of short sequences sequenced to the unique alignment on the X and Y chromosomes (N x , N y Calculating the ratio of the number of short sequences sequenced by the unique alignment to the short sequence number (M) of the whole genome unique alignment sequence, respectively, X-UR% and Y-UR%, when ⁇ -13 ⁇ 4% is greater than the first threshold When Y-UR% X-UR% is greater than the second threshold, it is determined that there is a male fetus in the twins.
  • the gender of the twins is determined by combining the fetal DNA concentration in the plasma of the pregnant woman. The specific method has been described in detail above and will not be described again.
  • the first threshold is 0.0004 (i.e., greater than 0.0004 is determined to be the presence of a male fetus) and the second threshold is 0.0075 (i.e., greater than 0.0075 is determined to be the presence of a male fetus).
  • the total number of sequencing data located on a chromosome is proportional to the length of the chromosome and the content of the chromosome in the biological sample
  • the sequencing data content of the Y chromosome in the sample is Y-UR%, or Y-UR% X-UR. / value, can effectively determine whether there are male fetuses in the twins. Further, based on the fetal DNA concentration in the plasma of the pregnant woman, the sex of the two fetuses in the twins can be further determined.
  • the present invention provides a computer readable medium.
  • the computer readable medium stores instructions thereon, the instructions being adapted to be executed by the processor to determine in the twins by the following steps Fetal gender:
  • the sequencing data is aligned with a human reference genomic sequence to obtain a unique aligned sequencing data set consisting of multiple unique aligned sequencing data.
  • the reference genomic sequence is a human genome reference sequence of version 36 of the NCBI database.
  • the sequencing data is aligned with the reference genomic sequence of the organism using at least one of SOAP and BWA.
  • the sequencing data has a length of 36 100 bp.
  • the number M of unique aligned sequencing data contained in the unique aligned sequencing data set is determined.
  • the unique alignment sequencing data sets at least one of the number N of unique alignment sequencing data derived from the Y chromosome and the number N x of unique alignment sequencing data derived from the X chromosome.
  • the sequencing data content of the Y chromosome is determined to be Y-UR 0 /.
  • determining the sex of the fetus in the twins wherein, when at least one of the following conditions is satisfied, determining that at least one of the twins is a male fetus, and determining that the twins are female Y when none of the following conditions are satisfied -UR% is greater than the first threshold; and ⁇ -1; 1% «-13 ⁇ 4% is greater than the second threshold.
  • the first threshold is 0.0004 and the second threshold is 0.0075.
  • the method when it is determined that at least one of the twins is a male fetus, the method further includes: performing fetal DNA concentration estimation based on the N x and ⁇ ⁇ , respectively, to obtain the first fetus separately DNA concentration. /. chrX, and second fetal DNA concentration % ⁇ 1; and determining the %chrX and %.
  • the computer-readable shield is used to determine the twin
  • the Y-UR%, or Y-UR%/X-UR% value of the Y chromosome in the blood DNA sample of the pregnant woman can effectively determine whether there is a male fetus in the twin. Further, based on the fetal DNA concentration in the maternal plasma, the sex of the two fetuses in the twins can be further determined.
  • the invention proposes a system for determining the sex of a fetus in a twin.
  • the system includes: a sequencing device A100, a comparison device A200, and an analysis device A300.
  • the sequencing device A100 is configured to sequence blood DNA of a twin pregnant woman to obtain a plurality of sequencing data
  • the comparison device A200 is connected to the sequencing device A100 for using the sequencing data with a human reference genome
  • the sequences are aligned to obtain a unique aligned sequencing data set consisting of a plurality of unique aligned sequencing data
  • analytical device A300 is coupled to comparison device A200 and is used to determine fetal gender in the twins.
  • the reference genomic sequence is the human genome reference sequence of version 36 of the NCBI database.
  • the sequencing data is aligned with the reference genomic sequence of the organism using at least one of SOAP and BWA.
  • the analyzing device A300 further includes: a first calculating module A310, configured to determine the number of unique aligned sequencing data included in the unique aligned sequencing data set! a second calculation module A320, configured to determine the number N y of unique alignment sequencing data derived from the Y chromosome in the unique alignment sequencing data set, and the number of unique alignment sequencing data derived from the X chromosome N x At least one of; a third calculating module A330, configured to determine a sequencing data content Y-UR of the Y chromosome.
  • the first threshold is 0.0004 and the second threshold is 0.0075.
  • Y-UR% is greater than 0.0004 or Y-UR%/X-UR% is greater than 0.0075, it can be determined that a male fetus is present in the twin.
  • the judging module A340 may further include a verification unit adapted to perform the following operations when determining that at least one of the twins is a male fetus: performing the fetus separately based on the N x and N Y
  • the DNA concentration is estimated to obtain a first fetal DNA concentration %chrX, and a second fetal DNA concentration %chrY, respectively; and to determine the % 1 «3 ⁇ 4 sum. /.
  • p-value of chrY wherein, if p-value>0.1, it is determined that the fetus in the twins is a male fetus, if p-value ⁇ 0.1, and %(;1 ⁇ is greater than %chrY, then determining There is one male and one female in the twins.
  • the system is used to determine the blood DNA of the pregnant woman of the twins.
  • the Y-UR%, or the value of Y-UR%/X-UR%, of the Y chromosome in the sample can be effectively confirmed. Whether there are male fetuses in twins. The sex of the two fetuses in the twins can then be effectively determined based on the fetal DNA concentration in the maternal plasma.
  • the present invention proposes another system for determining the sex of a fetus in a twin.
  • the system may include: a sequencing device for sequencing genomic DNA of the biological sample to obtain a plurality of sequencing data; and the computer readable medium described above.
  • the system is used to determine the Y chromosome in the blood DNA sample of the twin pregnant woman.
  • the sequencing data content of Y-UR%, or the value of Y-UR%/X-UR%, can effectively determine whether a male fetus is present in the twins. Further, based on the fetal DNA concentration in maternal plasma, the sex of the two fetuses in the twins can be effectively determined.
  • the method, system and computer readable medium of the present invention for determining the sex of a fetus in a twin have the following advantages:
  • the following steps are performed to verify the chromosome number of the patient.
  • Abnormal and twin gender conditions :
  • the DNA of the above 2 plasma samples (ie, 1 twin T21 positive sample and 1 twin fetal T18 positive sample, hereinafter referred to as Sl, S2) was extracted according to the Tiangen DP327-02 Kit protocol.
  • the extracted DNA was according to the modified Illumina/Solexa standard.
  • the library construction process is carried out, and the linker used for sequencing is added to both ends of the 200 bp DNA molecule.
  • Each sample is added with a different tag sequence (index), and then hybridized with the flowcell surface complementary link under certain conditions.
  • the nucleic acid molecules were clustered and then sequenced by single-end sequencing on IlluminaHiseq2000 to obtain a DNA fragment sequence of 36 bp in length.
  • DNA obtained from the above plasma sample was constructed according to the modified Illumina/Solexa standard procedure, and the specific procedure was referred to the prior art (see Illumina/ available at http://www.mumina.com7).
  • the Solexa standard library specification is incorporated by reference in its entirety.
  • the DNA library size and insert were determined to be about 200 bp by the 2100 Bioanalyzer (Agilent), and the QPCR was accurately quantified and used.
  • each sample was obtained with a data volume of about 0.25 G, wherein each sample was subjected to the aforementioned index label.
  • the districts are separated.
  • the obtained sequencing data of each sample is compared with the human genome reference sequence of version 36 (hgl8; NCBIBuild36) in the NCBI database, so as to obtain multiple unique comparisons of each sample respectively.
  • the unique alignment of the sequencing data consists of sequencing data sets.
  • the ratio of the average number of sequencing sequences ( n ' ) of the color bodies is 2, which is the base pair value, that is,
  • Threshold and Filtering - To obtain reliable results, filter the average normalized value of each chromosome. If the average normalized value is less than -1.28 or greater than 1.28 (95% CI), it will be output as a positive result.
  • Table 1 lists the karyotype results of the samples S1 and S2, Figure 3 shows the chromosomal digital karyotype results of the sample S1 obtained by the method of the present invention; and Figure 4 shows the sample S2 obtained by the method of the present invention. Chromosome digital karyotype results.
  • ratio of unique sequencing data were determined from the concentration of each sample X chromosome unique sequencing data than the number N x; statistics were unique to the ratio of the X chromosome and ⁇ of each sample For sequencing short sequence numbers N x , N y , where ⁇ is the number of unique alignment sequencing data derived from the X chromosome, and N y is the number of unique alignment sequencing data derived from the Y chromosome. Then, the ratio of the above-mentioned unique alignment sequencing short sequence number to the whole genome unique alignment sequencing short sequence number (M) was calculated separately, and respectively recorded as X-UR 0 /.
  • Y-UR% is greater than 0.0004, it is determined that there is a male fetus, Y-UR%/X-UR°/.
  • 0.0075 it is determined that there is a male fetus.
  • the sex of the two fetuses in the twins is determined. Specifically, %chrX with%. When 1 « ⁇ significant difference p-value>0.1, it can be inferred that the two fetuses are two male fetuses, when %chrX and %. 111" ⁇ significant difference - ⁇ 111 ⁇ 2 ⁇ 0.1) and %(;111 ratio. /. When chrY is high, it can be inferred that the two fetuses are a male fetus and a fetus.
  • the body of the twin fetus can be inferred by reference to the estimated fetal DNA concentration of the variant chromosome. Specifically, if the estimated fetal DNA concentration of the variant chromosome is about 1/2 of the estimated fetal DNA concentration of the Y chromosome, then the two fetuses are inferred. For the two male tires.
  • chromosomal variation eg, aneuploid
  • the body of the twin fetus can be inferred by reference to the estimated fetal DNA concentration of the variant chromosome. Specifically, if the estimated fetal DNA concentration of the variant chromosome is about 1/2 of the estimated fetal DNA concentration of the Y chromosome, then the two fetuses are inferred. For the two male tires.
  • the inventor concludes that at least one of the twins of the sample S1 is a male tire, and further, according to the front
  • the inference method because %chrX and %(;11 ⁇ estimated fetal DNA concentration difference is significant, the p-value is 0.408, so it can be determined that the twins are male and female.
  • the %chrX and % ⁇ differential significance p-value is 0.051
  • the estimated fetal DNA concentration (%chrY) based on the Y chromosome is 2.55 times the estimated fetal DNA concentration of the 18th variant chromosome, thereby determining that the S2 sample is two Male tires.
  • %chrX represents the fetal DNA concentration estimated by the relatively unique alignment of the short sequence number on the X chromosome
  • %chrY represents the fetal DNA concentration estimated by the relatively unique alignment of the short sequence number on the Y chromosome
  • %clirl8/%chr21 The fetal DNA concentration estimated by the relatively unique alignment of the sequenced short sequences on the chromosome that is mutated.
  • the method, system and computer readable medium of the present invention for determining the sex of a fetus in a twin can be effectively used to determine the sex of a fetus in a twin of a twin pregnant woman.

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Abstract

L'invention a trait à un procédé de détermination sexuelle d'embryons de jumeaux, à un système et à un support lisible par ordinateur connexes. Le procédé comprend une étape de séquençage de l'ADN sanguin d'une femme enceinte de jumeaux; une étape de comparaison des données de séquençage afin de collecter des données de comparaison unique; une étape de détermination d'un nombre M de données de séquençage recueillies parmi les données de séquençage; une étape de détermination d'au moins un nombre parmi Nx et Ny respectivement dérivés des données de séquençage de comparaison unique du chromosome X et du chromosome Y; une étape de détermination d'au moins un type de Y-UR en pourcent du chromosome Y et de X-UR en pourcent du chromosome X dans les données de séquençage; et une étape de détermination sexuelle d'embryons de jumeaux. Lorsqu'au moins un résultat remplit les conditions, au moins un des embryons de jumeaux est mâle, dans le cas contraire, les embryons sont tous féminins, à savoir le pourcentage de Y-UR dépasse une première valeur seuil et le rapport en pourcent Y-UR / X-UR dépasse une seconde valeur seuil.
PCT/CN2012/087582 2012-12-26 2012-12-26 Méthode de détermination sexuelle d'embryons de jumeaux, système et support lisible par ordinateur connexes WO2014101024A1 (fr)

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Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20120219950A1 (en) * 2011-02-28 2012-08-30 Arnold Oliphant Assay systems for detection of aneuploidy and sex determination

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20120219950A1 (en) * 2011-02-28 2012-08-30 Arnold Oliphant Assay systems for detection of aneuploidy and sex determination

Non-Patent Citations (2)

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Title
JUN WANG ET AL.: "The diploid genome sequence of an Asian individual", NATURE, vol. 456, no. 7218, 2008, pages 60 - 66 *
YUTAKA NAKAHORI ET AL.: "Sex Identification by Polymerase Chain Reaction Using X-Y Homologous Primer", AMERICAN JOURNAL OF MEDICAL GENETICS, vol. 39, no. 4, 1991, pages 472 - 473 *

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