CN117757798A - Hereditary microcephaly pathogenic mutant gene and application thereof - Google Patents

Abstract

The invention provides a novel mutant gene related to hereditary microcephaly and related application of the mutant gene. Specifically, the invention provides a mutant gene, which is c.8652dup mutant and/or c.8044C > T mutant in the coding region of wild-type ASPM gene compared with wild-type ASPM gene. Meanwhile, the invention further provides nucleic acid and polypeptide related to the mutant gene and application of the mutant gene, the nucleic acid or the polypeptide in preparation of a reagent, a kit or equipment for detecting the microcephaly. Meanwhile, the invention further discloses a kit for detecting the microcephaly. The new pathogenic mutation provided by the invention can be used for molecular diagnosis of patients with small head deformity and differential diagnosis of related diseases, is quick, accurate, efficient, simple and convenient, has high early diagnosis rate, and the detection result can provide scientific basis for early diagnosis, differential diagnosis and drug treatment of autosomal recessive primary small head deformity.

Description

Hereditary microcephaly pathogenic mutant gene and application thereof

Technical Field

The invention relates to the field of biology, in particular to a mutant gene related to hereditary microcephaly, application thereof and a related kit.

Background

The small head deformity is defined as the occipital forehead (occipitofrontal head circumference) being lower than 2 and more standard deviations (standard deviation, SD) of the average of the same age and sex newborns, where < the average of the head circumference-3 SD is referred to as severe small head deformity. The causes of the microcephaly are numerous, including genetic factors, environmental factors, viral infections, etc., with genetic factors being the most important factors. The microcephaly is classified into primary and secondary microcephaly according to the time of onset.

Primary small head deformities (primary microcephaly, PM) are a group of autosomal recessive or dominant diseases characterized by a slow down of brain development from fetal stage, a variable severity of intellectual disability (intellectual disability, ID), usually not involving extracentral nervous system deformities.

The etiology of the microcephaly directly affects the brain structure and growth of the infant, and can prevent the brain development of the infant in the prenatal period, perinatal period or postnatal period. At present, no effective drug treatment means is available for the primary microcephaly, so that the etiology of microcephaly is determined by a gene detection means, and prenatal gene diagnosis and intervention measures are further adopted, thus the method is an effective means for reducing the childbearing mental retardation.

ASPM is located at chromosome 1q31.3, contains 28 exons, and is autosomal recessive primary microcephaly type 5 caused by mutation, belonging to higher microcephaly in the population.

Disclosure of Invention

Through intensive research and analysis of ASPM genes, the invention provides a novel mutant gene related to hereditary microcephaly and related application of the mutant gene. Based on the genetic microcephaly pathogenic mutation which is not disclosed, the genetic microcephaly pathogenic mutation can be used as a candidate screening site of a microcephaly screening product to develop a diagnosis kit, a diagnosis chip and the like, and the data volume of a clinical mutation database is increased.

One aspect of the present invention discloses novel mutant genes associated with small head deformities that produce c.8652dup mutations and/or c.8044c > T mutations in the coding region of a wild-type ASPM gene as compared to the wild-type ASPM gene.

Specifically, the above-mentioned c.8652dup mutation refers to repeated insertion of one A base into the A base at position 8652 of exon18 of the wild-type ASPM gene (NM-018136.5). The c.8044C > T mutation refers to the mutation of the C base of the 8044 th exon18 of the wild type ASPM gene into a T base. Both of the above mutation sites were first discovered by the present invention.

Also, the present invention provides a nucleic acid having a c.8652dup mutation and/or a c.8044c > T mutation as compared to the wild-type ASPM gene.

In a second aspect, the invention discloses a polypeptide associated with a small head deformity, expressed from the nucleic acid described above.

In a third aspect, the invention discloses the use of a mutant gene, nucleic acid or polypeptide as described above in the preparation of a reagent, kit or device for detecting a micropittus deformity.

It will be appreciated that the mutant gene of the present application is used as a newly discovered pathogenic mutant gene related to the small head deformity, and a person skilled in the art may design a detection reagent for the mutant gene sequence of the present application according to the existing mutant gene detection method and technology, to form a kit or a special detection device, for example, design a corresponding detection primer, a fluorescent probe, a gene hybridization probe, a lock probe, etc. for the mutant gene sequence or the nucleic acid sequence of the present application according to the conventional method, to detect the small head deformity. Similarly, the polypeptide of the present invention is a novel substance, and the presence of the polypeptide is directly related to the microcephaly, so that a person skilled in the art can design a detection reagent for the polypeptide of the present application according to the existing protein or polypeptide detection method to form a kit or a special detection device.

In a fourth aspect of the present invention, a recombinant vector comprising the above mutant gene or nucleic acid is disclosed.

In a fifth aspect, the present invention discloses a recombinant cell comprising the recombinant vector described above.

It will be appreciated that for ease of investigation, the mutant genes of the invention may be inserted into vectors to make recombinant vectors for convenience of subsequent investigation or detection. The specific type of vector may be selected according to the purpose of the study, for example, the vector may be selected from conventional pMD18-T or pMD19-T, etc.; as a method for preparing the recombinant vector, reference is made to the prior art. Similarly, the recombinant cells are also obtained by introducing the recombinant vector into which the mutant gene of the present invention is inserted for the convenience of subsequent studies or detection, and the specific host cell type may be selected according to the purpose of the study, and is not limited herein.

The sixth aspect of the invention discloses application of the mutant gene, the nucleic acid, the polypeptide, the recombinant vector or the recombinant cell in preparing a medicament for treating the microcephaly.

It will be appreciated that based on the above mutant genes, nucleic acids, polypeptides, recombinant vectors or recombinant cells disclosed herein, one skilled in the art can modify the relevant mutation sites for appropriate drugs to which the mutant genes are designed to return to a normal wild-type state or other state so as not to cause disease. Meanwhile, when a proper biological model is needed to be adopted in the preparation of the medicine for treating the microcephaly, the mutant genes, the nucleic acid, the polypeptide, the recombinant vector or the recombinant cell can be used as a research object (biological model per se) or can be prepared into other proper biological models (such as animal models) through the mutant genes, the nucleic acid, the polypeptide, the recombinant vector or the recombinant cell.

The seventh aspect of the invention also discloses a kit for detecting the microcephaly, which comprises a reagent for detecting the mutant gene or the nucleic acid and/or a reagent for detecting the polypeptide.

In one embodiment of the present invention, the above-mentioned reagents include probes and/or primers for specifically detecting the above-mentioned mutant genes or nucleic acids of the present invention. Specifically, the primers include the following amplification primers:

primer pair 1:

the upstream primer ASPM-F1, SEQ ID No.1:5'-CACAAGAAAACTGGAAACACA-3'; the downstream primer ASPM-R1, SEQ ID No.2:5'-TGCTGTCTTGGCTGACTTAC-3';

primer pair 2:

the upstream primer ASPM-F2, SEQ ID No.3:5'-CAGATTCAGGAACAGCACCA';

the downstream primer ASPM-R2, SEQ ID No.4:5'-AAGCAAGGCCAGAAGCTTTA-3'.

The invention has the beneficial effects that: the invention discovers 2 new pathogenic mutations of autosomal recessive primary small head deformity 5-type pathogenic gene ASPM, the mutation site can be used for molecular diagnosis of small head deformity patients and differential diagnosis of related diseases, and the detection result can provide scientific basis for early diagnosis, differential diagnosis and drug treatment of autosomal recessive primary small head deformity, and is rapid, accurate, efficient, simple and convenient, and high in early diagnosis rate.

Drawings

FIG. 1 is a schematic diagram of an autosomal recessive primary microcephaly Trio family of example 1, the family comprising 3 members, son being microcephaly patients (i.e., II-1 in the family diagram), parents being normal (i.e., I-1, I-2 in the family diagram), wherein solid icons are patients, and arrows indicate forerunner;

fig. 2 is a cranial MRI result for a patient in the family of example 1, wherein, graph a: craniocerebral MRI axis T2WI, panel B: the result of the brain MRI disfigurement T1WI shows that the brain width of the frontal lobe of the two sides is big, the sulcus is reduced, the brain is shallowed, the grey white matter is used for prompting the megabrain return deformity;

FIG. 3 is a diagram of representative Sanger sequencing verification peaks for ASPM gene c.8652dup mutation by members of family, patient II-1 carrying the c.8652dup mutation of ASPM gene, patient's mother I-2 carrying the c.8652dup mutation, patient I-1's father not carrying the c.8652dup mutation;

FIG. 4 is a diagram of representative Sanger sequencing verification peaks of the ASPM gene c.8044C > T mutation for each member of the family, patient II-1 carries the c.8044C > T mutation, patient I-1's father carries the c.8044C > T mutation, and patient's mother I-2 does not carry the c.8044C > T mutation.

Detailed Description

Full exon sequencing (WES) is the most frequently used method of sequencing genomes. Exons are protein-coding regions of the human genome whose DNA can be captured and enriched using sequence capture techniques. Although the exon region accounts for only about 1% of the whole genome, 85% of pathogenic mutations are contained. Compared with whole genome sequencing, whole exon sequencing is more economical and efficient. Exome sequencing is mainly used to identify and study mutations in coding regions and UTR regions associated with disease, population evolution. The combination of the exon data provided by the large public databases is advantageous for better interpretation of the resulting mutation versus disease. Thus, the inventors performed pathogenic variation detection and verification by a method combining whole exon sequencing and family analysis with Sanger sequencing verification for one self-collected small-head malformed Trio family (parent+forerunner). Finally, based on various detection results, the inventors determined 2 new pathogenic mutation sites, namely the c.8652dup mutation and the c.8044C > T mutation of the ASPM gene (NM-018136.5), resulting in autosomal recessive primary Microcephaly type 5 (Microcephaly 5,primary,autosomal recessive,MCPH5,OMIM: # 608716). The mutant genes are not reported in the literature at present.

Here, the DNA sequence (e.g., intron sequence, exon sequence, etc.), RNA sequence, encoded protein information, etc. of the wild-type ASPM gene are all recorded in the NCBI database, and reference is made to the following website https:// www.ncbi.nlm.nih.gov/gene/259266.

For ease of illustration, the NM-018136.5:c.8652dup (NG-015867.1:g.51096dup) mutation, a partial DNA sequence of the wild type ASPM genome (hg 19 human reference genome, reference sequence: NG-015867.1) is selected, the sequence being located at position 50931-51449 of the genome as shown in SEQ ID No. 5.

SEQ ID No.5 (sequence segment: EXON18-Intron18, base when not mutated, underlined):

CACAAGAAAACTGGAAACACAGAAATGTGCTGCCCTACGGATTCAGTTCTTCCTTCAGATGGCTGTGTATCGGAGAAGATTTGTTCAGCAGAAAAGAGCTGCTATCACTTTACAGCATTATTTTAGGACGTGGCAAACCAGAAAACAGTTTTTACTATATAGAAAAGCAGCAGTGGTTTTACAAAATCACTACAGAGCATTTCTGTCTGCAAAACATCAAAGACAAGTCTATTTACAGATCAGAAGCAGTGTTATCATTATTCAAGCTAGAAGTAAAGGATTTATACAGAAACGGAAGTTTCAGGAAATTAAAAATAGCACCATAAAAATTCAGGTATTTCTATATTTAATTTAAATATAAAACTGTGAGTCTTTCTTTTGACCAATATTTGTTAGAATGTATAAAATGAGCTTAAGCAGGTCATAAAGAATAGTACTTAGTTGAGGTGACCATTTATCTTCCAAAGTGGAACATTTTTGAAAGAGGGAAAGGGCACTAGTAAGTCAGCCAAGACAGCA

the c.8652dup mutation is defined herein by reference to the above sequence. The partial DNA of the mutant gene of the present invention, which corresponds to the partial DNA sequence of the wild-type ASPM gene (hg 19 human reference genome, reference sequence: NG_ 015867.1), is shown in SEQ ID No.6, and specifically, the c.8652dup mutation means that one A base is repeatedly inserted into the A base at position 8652 of exon18 of the wild-type ASPM gene.

SEQ ID No.6 (mutation sites are underlined, showing bases after mutation): CACAAGAAAACTGGAAACACAGAAATGTGCTGCCCTACGGATTCAGTTCTTCCTTCAGATGGCTGTGTATCGGAGAAGATTTGTTCAGCAGAAAAGAGCTGCTATCACTTTACAGCATTATTTTAGGACGTGGCAAACCAGAAAACAGTTTTTACTATATAGAAAAAGCAGCAGTGGTTTTACAAAATCACTACAGAGCATTTCTGTCTGCAAAACATCAAAGACAAGTCTATTTACAGATCAGAAGCAGTGTTATCATTATTCAAGCTAGAAGTAAAGGATTTATACAGAAACGGAAGTTTCAGGAAATTAAAAATAGCACCATAAAAATTCAGGTATTTCTATATTTAATTTAAATATAAAACTGTGAGTCTTTCTTTTGACCAATATTTGTTAGAATGTATAAAATGAGCTTAAGCAGGTCATAAAGAATAGTACTTAGTTGAGGTGACCATTTATCTTCCAAAGTGGAACATTTTTGAAAGAGGGAAAGGGCACTAGTAAGTCAGCCAAGACAGCA

To facilitate the description of the c.8044C > T mutation, the partial DNA sequence of the wild-type ASPM gene (hg 19 human reference genome, reference sequence: NM-018136.5) is selected herein, and the sequence is located from position 7858 to position 8413 of the coding region of the gene, as shown in SEQ ID No. 7.

SEQ ID No.7 (sequence segment: EX)ON18, underlined is the base when not mutated): CAGATTCAGGAACAGCACCAGGCTGCCATTATTATTCAGAAGCATTGTAAAGCCTTTAAAATAAGGAAGCATTATCTCCACCTTAGAGCAACAGTAGTTTCTATTCAAAGAAGATACAGAAAACTAACTGCAGTGCGTACCCAAGCAGTTATTTGTATACAGTCTTATTACAGAGGCTTTAAAGTACGAAAGGATATTCAAAATATGCACCGGGCTGCCACACTAATTCAGTCATTCTATCGAATGCACAGGGCCAAAGTTGATTATGAAACAAAGAAAACTGCAATTGTGGTTATACAGAATTATTATAGGTTGTATGTTAGAGTAAAAACAGAAAGAAAAAACTTTTTAGCAGTTCAGAAATCTGTACGAACTATTCAGGCTGCTTTTAGAGGCATGAAAGTTAGACAAAAATTGAAAAATGTATCAGAGGAAAAGATGGCAGCCATTGTTAACCAATCTGCACTCTGCTGTTACAGAAGTAAAACTCAGTATGAAGCTGTTCAAAGTGAAGGTGTTATGATTCAAGAGTGGTATAAAGCTTCTGGCCTTGCTT

Herein, the above c.8044c > T mutation is determined with reference to the above sequence. Corresponding to the partial DNA sequence of the wild-type ASPM gene, the partial DNA of the mutant gene of the present invention is shown in SEQ ID No.8, and specifically, the c.8044C > T mutation means that the C base of the 8044 th exon18 of the wild-type ASPM gene is mutated into a T base.

SEQ ID No.8 (mutation sites are underlined, showing bases after mutation): CAGATTCAGGAACAGCACCAGGCTGCCATTATTATTCAGAAGCATTGTAAAGCCTTTAAAATAAGGAAGCATTATCTCCACCTTAGAGCAACAGTAGTTTCTATTCAAAGAAGATACAGAAAACTAACTGCAGTGCGTACCCAAGCAGTTATTTGTATACAGTCTTATTACAGAGGCTTTAAAGTATGAAAGGATATTCAAAATATGCACCGGGCTGCCACACTAATTCAGTCATTCTATCGAATGCACAGGGCCAAAGTTGATTATGAAACAAAGAAAACTGCAATTGTGGTTATACAGAATTATTATAGGTTGTATGTTAGAGTAAAAACAGAAAGAAAAAACTTTTTAGCAGTTCAGAAATCTGTACGAACTATTCAGGCTGCTTTTAGAGGCATGAAAGTTAGACAAAAATTGAAAAATGTATCAGAGGAAAAGATGGCAGCCATTGTTAACCAATCTGCACTCTGCTGTTACAGAAGTAAAACTCAGTATGAAGCTGTTCAAAGTGAAGGTGTTATGATTCAAGAGTGGTATAAAGCTTCTGGCCTTGCTT

It is understood that one allele is a c.8652dup mutation and the other allele is a c.8044c > T mutation, resulting in the occurrence of autosomal recessive microcephaly. When carrying the c.8652dup mutation alone or the c.8044c > T mutation alone, it did not lead to the occurrence of autosomal recessive microcephaly, but was merely the carrier of the pathogenic mutation, the parent of the patient in the study. Based on the above-mentioned results, in practical applications, the two mutation sites can be detected simultaneously, for example, the mutation (only c.8652dup or only c.8044c > T mutation) is detected, and the subject may not develop the symptoms of the microcephaly related to the mutation, but the result suggests that the subject carries inheritable recessive pathogenic mutation. When detecting that the parents carry one of the two mutations (the same or different), the offspring of the parents can be simultaneously inherited to the recessive pathogenic mutation to be diseased.

Regarding the practical application of the mutation sites, in one embodiment of the present invention, two pairs of primers (primer pair 1 and primer pair 2, as shown in SEQ ID No. 1-4) are designed and used to specifically amplify the DNA of the sample to be detected, respectively, so as to realize the separate detection of the two mutation sites. Furthermore, the mutant gene detection of the present invention can also be detected together with other mutant genes related to the microcephaly, namely, the gene detection panel is formed. Therefore, the design of the detection primer or probe and the selection of the specific detection method can be adjusted according to the actual requirements of the detection, and do not limit the present invention.

It will be appreciated that the reagents for detecting the mutant gene sequences of the present invention are not limited to the primers shown in SEQ ID Nos. 1 to 4, and that more amplification primers may be designed for the c.8652dup and c.8044C > T mutations, where the amplification primers specifically refer to primers in which the amplified fragment contains the c.8652dup mutation and/or the c.8044C > T mutation, and after DNA is amplified by these amplification primers, whether the amplified fragment contains the mutation of the present invention is analyzed, thereby confirming the presence or absence of the mutant gene sequences of the present invention. In addition, specific primers or specific probes, such as real-time fluorescent probes, gene hybridization probes, lock probes, etc., can also be designed based on the c.8652dup mutation and the c.8044c > T mutation; the method for confirming the c.8652dup mutation and the c.8044C mutation is not limited to sequencing, and can also confirm the c.8652dup mutation and the c.8044C mutation through specific primers or specific probes; sequencing is, of course, the most direct, efficient and accurate method.

The invention will be described in further detail below with reference to the drawings by means of specific embodiments.

Example 1 determination of autosomal recessive primary kohlrabi pathogenic mutations

1. Sample collection

The inventor collects Trio family (parent+forensics) of a Chinese Han nationality small-head malformation patient, and the family diagram is shown in fig. 1. As shown in FIG. 1, the family contains 3 members, the son is a person with a small head deformity (i.e. II-1 in the family chart) and the parent is a normal person (i.e. I-1, I-2 in the family chart). Wherein, the solid icons are patients, and the arrows are forerunner.

First-person pregnancy at fetal stage 23 ⁺⁶ W ultrasound, bpd=51.6 mm (-3.5 SD), hc=193 mm (-3 SD); pregnancy 31 ⁺³ W bpd=70 mm (-3.9 SD), hc=249 mm (-4.3 SD); pregnancy 39 ⁺² W delivery following delivery A baby boy had a 1min Apgar score of 10 minutes, no history of oxygen deficiency, a body weight of 2700g, a length of 47cm, HC=29 cm (-3.4 SD).

The results of the skull MRI of the patients in this family are shown in fig. 2. In fig. 2, a: craniocerebral MRI axial position T2WI; b: craniocerebral MRI disfigurement T1 WI), the results show bilateral frontal parietal gyrus width, reduced sulcus, pale grey matter, suggesting giant gyrus deformity.

The inventor collects peripheral blood samples of all members in the family, and EDTA is added for anticoagulation and preservation at-80 ℃. All blood samples were signed with informed consent.

2. DNA extraction

Peripheral blood of all members of the family is taken, genomic DNA of peripheral blood leucocytes is extracted by QIAmp blood kit (Qiagen, hilden, germany) respectively, and the concentration and purity of the DNA are measured by using Qubit Fluorometer and agarose gel electrophoresis, and the obtained genomic DNA OD260/OD280 of each specimen is between 1.7 and 2.0, and the concentration is not less than 50 nanograms per microliter, and the total amount is not less than 3 micrograms.

3. Capture sequencing

Samples of all family members were sequenced using the KAPA HyperExome et al capture system in combination with the MGISEQ-2000 high throughput sequencing technique. The capture range includes whole genome exon regions (about 1% of whole genome), the total length of the capture region can reach 50M, and the sequencing data volume is 10-12Gb, and the effective sequencing depth is 100 times. The main steps include disruption, library preparation and on-machine sequencing.

After the sequencing data is taken off the machine, mutation detection, annotation and database comparison are carried out by using an internal customization flow, and candidate pathogenic sites are determined according to means such as crowd frequency, software prediction results, family analysis and the like.

Results: 2 heterozygous mutations were found on the ASPM gene of the patient, c.8652dup and c.8044c > T, respectively. Family analysis shows that the mother of the patient carries a c.8652dup mutation and the father of the patient carries a c.8044c > T mutation, suggesting that 2 mutations are complex heterozygous.

If the c.8652dup and c.8044C > T mutations were positive as true as verified by Sanger, it was essentially confirmed that the mutation was causative of the patient's micropitter.

Example 2Sanger sequencing validation

ASPM genes of all family members in the family of the micropittic patient described in example 1 were examined separately: primers are designed for the mutations c.8652dup and c.8044C > T of the ASPM gene, then related sequences of mutation sites are obtained through the methods of PCR amplification, product purification and sequencing, and whether the mutations c.8652dup and c.8044C > T of the ASPM gene are detected in a sample is verified according to the determination result of determining whether the sequences belong to a mutant type or a wild type.

The method comprises the following specific steps:

1. DNA extraction

Genomic DNA from peripheral venous blood of a subject was extracted for use according to the DNA extraction method described in example 1.

2. Primer design and PCR reaction

First, referring to the human genome reference sequence GRCh37/hg19, specific primers were designed for the c.8652dup and c.8044c > T mutations of ASPM genes, the specific sequences were as follows:

TABLE 1Sanger sequencing primers

Then, a PCR reaction system for each DNA sample was prepared in accordance with the proportions shown in Table 2 below, and a PCR reaction was performed in accordance with the reaction conditions shown in Table 3.

TABLE 2PCR reaction system (25 μl)

TABLE 3PCR reaction conditions

Thus, PCR amplification products of the genomic DNA samples of the respective subjects were obtained.

3. Sanger sequencing

And (3) purifying the PCR product obtained in the step (2), directly performing DNA sequencing, and performing forward and reverse sequencing by using an ABI3730XL type sequencer.

Based on the sequencing results, as shown in fig. 3 and 4, in the shown patient Trio family, the patient carried ASPM gene c.8652dup and c.8044c > T mutations, the normal mother carried c.8652dup mutations, and the normal father carried c.8044c > T mutations, suggesting that 2 mutations were complex heterozygous.

In combination with the above information, it was confirmed that the c.8652dup and c.8044C > T mutations of the ASPM gene are causative agents of the II-1 microcephaly of patients.

It was further found by investigation that the c.8652dup mutation caused in-frame coding due to insertion of base a, resulting in mutation of alanine at amino acid 2885 to serine, resulting in premature stop codon at 2920 (a 2885 sfs.35). The 8044C > T variant mutates the 8044 nucleotide from C to T, resulting in nonsense mutation of arginine at 2682 (R2682X). Both variants are predicted to lead to premature truncation of ASPM proteins, lack of partial IQ repeat regions and Armadillo-like regions, and thus to disease development.

Example 3 detection kit

A detection kit was prepared comprising primers capable of detecting the c.8652dup and c.8044c > T mutation of the ASPM gene for diagnosing biological samples of autosomal recessive primary micropitta, wherein these primers are ASPM gene specific primers having the sequences shown in table 1 of example 2 (SEQ ID nos. 1-4).

The specific steps of diagnosing autosomal recessive primary microcephaly biological sample by using the kit are as follows:

the DNA of the subject is extracted according to the method described in step 2 of example 1, PCR reaction is performed with the specific primer of the above ASPM gene using the extracted DNA as a template (the PCR reaction system and reaction conditions are described in example 2), the PCR product is purified according to the conventional method in the art, the purified product is sequenced, and then whether the ASPM gene mutant of the present invention exists in the DNA of the subject can be effectively detected by observing whether the sequence obtained by sequencing has c.8652dup and c.8044C > T mutation, thereby being able to effectively detect whether the subject suffers from autosomal recessive primary microposity.

The foregoing description of the invention has been presented for purposes of illustration and description, and is not intended to be limiting. Several simple deductions, modifications or substitutions may also be made by a person skilled in the art to which the invention pertains, based on the idea of the invention.

Claims (10)

1. A mutant gene, wherein the mutant gene has a c.8652dup mutation and/or a c.8044c > T mutation from the coding region of a wild-type ASPM gene.

2. A nucleic acid, characterized in that the nucleic acid has a c.8652dup mutation and/or a c.8044c > T mutation compared to the wild-type ASPM gene.

3. A polypeptide expressed from the nucleic acid of claim 2.

4. Use of a mutant gene according to claim 1, a nucleic acid according to claim 2 or a polypeptide according to claim 3 for the preparation of a detection reagent, kit or device associated with a microcephaly.

5. A recombinant vector comprising the mutant gene of claim 1 or the nucleic acid of claim 2.

6. A recombinant cell comprising the recombinant vector of claim 5.

7. Use of a mutant gene according to claim 1, a nucleic acid according to claim 2, a polypeptide according to claim 3, a recombinant vector according to claim 5 or a recombinant cell according to claim 6 for the preparation of a medicament for the treatment of a microcephaly.

8. A kit for detecting a microcephaly, characterized in that: the kit comprises a reagent for detecting the mutant gene according to claim 1 or the nucleic acid according to claim 2, and/or a reagent for detecting the polypeptide according to claim 3.

9. The kit of claim 8, wherein: the reagents include probes and/or primers that specifically detect the mutant gene or nucleic acid.

10. The kit of claim 9, wherein: the primers include at least one of amplification specific amplification primer pairs, wherein,

primer pair 1:

the upstream primer ASPM-F1, SEQ ID No.1:5'-CACAAGAAAACTGGAAACACA-3'; the downstream primer ASPM-R1, SEQ ID No.2:5'-TGCTGTCTTGGCTGACTTAC-3';

primer pair 2:

the upstream primer ASPM-F2, SEQ ID No.3:5'-CAGATTCAGGAACAGCACCA';

the downstream primer ASPM-R2, SEQ ID No.4:5'-AAGCAAGGCCAGAAGCTTTA-3'.