Screening method and application of high-specificity chromosome probe combination

Abstract

The invention discloses a screening method and application of a high-specificity chromosome probe combination. Comprising the following steps: intercepting a sequence from a centromere region and / or a telomere region of a target chromosome, generating probes, comparing all the generated probes with the centromere region of the target chromosome, and screening out a probe with high specificity with the target chromosome; comparing the probe with a non-target chromosome, and screening out a probe with low specificity with the non-target chromosome; evaluating the stability of the probe, and screening out the probe with high stability; and screening according to the repetition times of the probe in the centromere areas of the target chromosome and the non-target chromosome, and after screening, outputting a finally screened high-specificity chromosome probe sequence. According to the screening method disclosed by the invention, the specific probe can be rapidly and efficiently screened in the specific region of the chromosome, the secondary structure of the probe is considered, and the specificity and the sensitivity of the probe are considered.

Description

Technical Field

[0001] The present invention relates to the technical fields of bioinformatics and genomics, and particularly relates to a method for screening a high-specificity chromosome probe combination and an application thereof. Background Art

[0002] With the development of high-throughput sequencing technology, more and more genomes have been sequenced and assembled completely. In genomics and molecular biology research, it is often necessary to face a huge sequence to find a specific region sequence and design more specific and accurate probes to detect specific region fragments on chromosomes.

[0003] Currently, when screening for sequences in a specific region within the chromosome scale, manual screening is mainly carried out through sequence alignment tools (such as BLAST, etc.), lacking a fast and efficient screening and design program, especially for some regions with highly repetitive sequences on chromosomes, such as centromere, telomere regions, etc. The existing methods and tools currently have obvious deficiencies in screening specific probes, which are mainly manifested in the following aspects:

[0004] 1. Insufficient probe specificity: Existing probe screening tools cannot effectively distinguish similar sequences on target chromosomes and non-target chromosomes, resulting in the probe being able to bind on non-target chromosomes, thus affecting the accuracy of the detection result.

[0005] 2. Non-Fold space structure (non-folding structure with complicated secondarystructure DNA domain) not considered: In probe screening, the existing technology usually ignores the spatial structure stability of the probe. When the probe binds to the target sequence, if it forms a Fold structure itself, it will significantly reduce its binding efficiency to the target, thereby affecting the sensitivity and specificity of the probe.

[0006] 3. Low screening efficiency: The current tool program has a low efficiency in screening probes multiple times and cannot generate high-specificity probes in a short time.

[0007] Therefore, there is an urgent need for an algorithm and tool that can simultaneously improve probe specificity, optimize probe structure, and improve screening efficiency to meet the urgent need for high-specificity probes in current genomic detection, and there is also an urgent need for a high-specificity chromosome probe combination. Summary of the Invention

[0008] The purpose of the present invention is to overcome the deficiencies of the prior art and provide a screening method and application of a high-specificity chromosome probe combination. The high-specificity chromosome probe combination of the present invention has high specificity and sensitivity. The screening method of the present invention can quickly and efficiently screen out specific probes in specific regions of chromosomes, and considers the secondary structure of the probe, taking into account the specificity and sensitivity of the probe.

[0009] To achieve the above purpose, the technical solution designed by the present invention is as follows:

[0010] The present invention provides a method for screening a high-specificity chromosome probe combination, comprising the following steps:

[0011] (1) Extract the sequence from the centromere region and / or telomere region of the target chromosome to generate probe P i The collection is the probe set;

[0012] (2) Each probe P in the probe set i Compare with the centromere region and / or telomere region of the target chromosome, select probes whose similarity with the target chromosome is greater than or equal to the threshold θ1, and select the probe P i-1 The set is the preliminary probe screening set;

[0013] (3) Each probe P in the preliminary probe screening set i-1 Compare with non-target chromosomes and select probes whose similarity with non-target chromosomes is less than or equal to threshold θ2; the probes P obtained by screening i-2 The set is the secondary probe screening set;

[0014] (4) Each probe P in the secondary probe screening set i-2 The stability of the probes was evaluated by using a non-fold structure prediction tool, and the probes with stability greater than or equal to the threshold θ3 were screened. i-3 The set is the stability probe screening set;

[0015] (5) Each probe P in the stability probe screening set i-3 The number of repetitions in the centromere region and / or telomere region of the target chromosome and the non-target chromosome is screened, and the number of repetitions of the screening probes in the target chromosome is greater than or equal to the number threshold r1, and the number of repetitions in the non-target chromosome is less than or equal to the number threshold r2. The screened probe P i-4 The set is the probe screening set for the number of repetitions;

[0016] (6) Finally, output the number of repetitions for each probe P in the probe screening set. i-4 , and obtain highly specific chromosome probe sequences.

[0017] Further, in the step (1), each probe P in the probe set i is designed according to the following formula:

[0018] P i = S i [start:end];

[0019] wherein, P i is a probe, i = 1, 2,..., n, S is the target chromosome sequence, [start, end] are the start and end positions of the centromeric region or telomeric region of the target chromosome, and n is the number of generated probes;

[0020] In the step (2), the screening conditions for each probe P in the preliminary probe screening set i-1 are as follows:

[0021] S target (P i ) ≥ θ1,

[0022] wherein, S target (P i ) is the similarity between the probe P i and the target chromosome, and θ1 is the similarity threshold between the probe P i and the target chromosome;

[0023] In the step (3), the screening conditions for each probe P in the secondary probe screening set i-2 are as follows:

[0024] S non-target (P i-1 ) ≤ θ2,

[0025] wherein, S non-target (P i-1 ) is the similarity between the probe P i-1 and non-target chromosomes, and θ2 is the similarity threshold between the probe P i-1 and non-target chromosomes;

[0026] In the step (4), the screening conditions for each probe P in the stability probe screening set i-3 are as follows:

[0027] F(P i-2 ) ≥ θ3,

[0028] wherein, F(P i-2 ) is the non-fold structure stability score of the probe P i-2 , and θ3 is the stability score threshold of the non-fold structure of the probe P i-2 ;

[0029] In the said step (5), each probe P in the probe screening set for the number of repetitions is repeated i-4 The screening conditions are as follows:

[0030] R target (P i-3 ) ≥ r1 and R non-target (P i-3 ) ≤ r2,

[0031] wherein, R target (P i-3 ) is the number of repetitions of probe P i-3 in the centromeric region and / or telomeric region of the target chromosome, and R non-target (P i-3 ) is the number of repetitions of probe P i-3 in the centromeric region and / or telomeric region of non-target chromosomes;

[0032] r1 is the threshold value of the number of repetitions of probe P i-3 in the centromeric region and / or telomeric region of the target chromosome, and r2 is the threshold value of the number of repetitions of probe P i-3 in the centromeric region and / or telomeric region of non-target chromosomes.

[0033] Furthermore, the non-fold structure prediction tool is NUPACK.

[0034] Furthermore, in the said step (2), the similarity threshold θ1 between probe P i and the target chromosome is 0.8;

[0035] In step (3), the similarity threshold θ2 between probe P i-1 and non-target chromosomes is 0.3;

[0036] In step (4), the stability score threshold θ3 of the non-fold structure of probe P i-2 is 0.7;

[0037] In step (5), the threshold value r1 of the number of repetitions of probe P i-3 in the centromeric region and / or telomeric region of the target chromosome is 5, and the threshold value of the number of repetitions of probe P i-3 in the centromeric region and / or telomeric region of non-target chromosomes is 1.

[0038] The present invention also provides an application of the said screening method in screening chromosome probes with high specificity and high sensitivity.

[0039] The present invention also provides a highly specific chromosomal probe combination screened by using the above method. The probe combination includes 10 probe sequences, namely Chr1_1, Chr1_2, Chr1_3, Chr1_4, Chr1_5, Chr1_6, Chr1_7, Chr1_8, Chr1_9 and Chr1_10, and their nucleotide sequences are shown in SEQ ID NO: 1 to SEQ ID NO: 10 respectively.

[0040] Further, the chromosome is human chromosome 1.

[0041] Still further, the probe combination is screened from the centromeric region of human chromosome 1.

[0042] The present invention also provides an application of the above probe combination in the preparation of a kit.

[0043] The present invention also provides a kit, which includes the above probe combination.

[0044] The present invention also provides an application of the above kit in the detection of human cells.

[0045] Advantages of the present invention:

[0046] The present invention provides an accurate and efficient method for screening probe sequences, which can target specific chromosomal position regions, especially repetitive regions such as centromeres and telomeres, and can screen out highly specific and efficient detection probes according to requirements to meet the research needs such as in situ hybridization (FISH). The algorithm proposed by the present invention effectively reduces the interference of similar sequences on non-target chromosomes, can provide high-quality probe design, and provides important tool support for genomic research and clinical applications. The highly specific chromosomal probe combination screened by using the screening method of the present invention has high specificity and sensitivity. Brief Description of the Drawings

[0047] Figure 1 It is a hybridization test result diagram of the highly specific chromosomal probe combination on human cells. Detailed Embodiments

[0048] The following further describes the present invention in detail with reference to specific embodiments for those skilled in the art to understand.

[0049] Example 1

[0050] A method for screening highly specific chromosomal probes

[0051] This embodiment relies on the Linux operating system and the Python programming language. In the Linux system, the present invention uses the BLAST tool for sequence alignment, and at the same time generates and screens probes through Python scripts.

[0052] I. Environmental requirements for the screening method:

[0053] 1. Operating system: Linux (Ubuntu 18.04 or later versions);

[0054] 2. Programming language: Python 3.6 and above versions;

[0055] 3. Required tools: BLAST+ 2.9.0 or higher versions;

[0056] 4. Other dependencies: Biopython, Numpy, Scipy;

[0057] 5. Hardware resources: Use a server environment with at least 16GB of memory and a 4-core processor for large-scale probe screening.

[0058] II. Input files for the screening method:

[0059] The input file of this system is the whole genome data corresponding to the target site, including the genomic sequence of the chromosome where the target site is located (target chromosome), and the genomic sequence library of all the remaining chromosomes (non-target chromosomes) (for example, if the target chromosome is human chromosome 1, the non-target chromosomes are the sum of the 23 pairs of chromosomes other than chromosome 1 in the human chromosomes), which is used for two rounds of alignment. The file format requirement is FASTA format.

[0060] III. Required computing resources for the screening method: According to the large-scale probe screening requirements of this system, it is recommended to use a server environment with the following configurations:

[0061] 1. Processor: 4 cores and above;

[0062] 2. Memory: At least 16GB;

[0063] 3. Hard disk: At least 500GB of available storage space for storing the alignment database and intermediate files.

[0064] IV. The screening method for highly specific chromosome probes mainly includes four contents: probe generation, alignment screening, structure prediction, and repeat screening, which specifically include the following steps:

[0065] 1. Probe generation. In the probe generation module, sequences are intercepted from the centromere and telomere regions of the target chromosome (according to actual needs, sequence interception can be performed only for the centromere or telomere region), and probes are generated. The generated probes can be expressed as:

[0066] P = S[start:end]

[0067] where P is the fragment intercepted from the chromosome sequence S and serves as the probe, S is the target chromosome sequence, and [start, end] are the start and end positions of the centromere region or telomere region of the target chromosome. (According to the provided file, the start and end coordinates of the chromosome can be set as (example): P = S[122026460 - 125184587], for example, the start position of the centromere of human chromosome 1);

[0068] For the case of generating multiple probes, a probe set is generated. Each probe P in the probe set i is designed by the following formula:

[0069] P i = S i [start:end];

[0070] In the formula, P i is the probe, i = 1, 2,..., n, S is the target chromosome sequence, [start, end] are the start and end positions of the centromere region or telomere region of the target chromosome, and n is the number of generated probes;

[0071] (1) Steps for the computer to execute:

[0072] a. In the Linux system, first install the required dependency packages through the following commands:

[0073] sudo apt-get update;

[0074] sudo apt-get install ncbi-blast+python3-pip;

[0075] pip3 install biopython numpy scipy.

[0076] b. Use a Python script to generate the preliminary probe sequence:

[0077] python3 IndexCreator.py --input target_chromosome.fasta --outputprobes.fasta.

[0078] 2. First-round comparison and screening. After the probes are generated, use the BLAST tool to compare each probe P in all the generated probe sets i with the centromere and telomere regions of the target chromosome.

[0079] Select the probes with a similarity S target (P i ) greater than or equal to the threshold θ1. The set of the selected probes P i-1 is the preliminary probe screening set. The preliminary probe screening conditions are as follows:

[0080] S target (P i ) ≥ θ1,

[0081] where S target (P i ) is the similarity between the probe P i and the target chromosome;

[0082] θ1 is the similarity threshold of the probe P i to the target chromosome;

[0083] Set the similarity threshold θ1 to 0.8, that is, it is required that the alignment similarity between the probe P i and the target chromosome is greater than or equal to 80%.

[0084] (1) The computer executes the steps: Use the BLAST tool for the first-round comparison and screening, and compare the generated probe sequences with the target chromosome:

[0085] blastn - query probes.fasta - db target_chromosome_db - out results_target.txt;

[0086] 3. Second-round comparison and screening. In the second-round comparison and screening, compare each probe P in the preliminary probe screening set i-1 with the non-target chromosomes, eliminate the probes that also have a high similarity on the non-target chromosomes, and select the probes with a similarity S non-target (P i-1 ) less than or equal to the threshold θ2; The set of the selected probes P i-2 is the secondary probe screening set. The secondary probe screening conditions are as follows:

[0087] S non-target (P i-1 ) ≤ θ2,

[0088] where S non-target (P i-1 ) is the similarity between the probe P i-1Similarity with non-target chromosomes;

[0089] θ2 is probe P i-1 Similarity threshold with non-target chromosomes; Set probe P i-1 The similarity threshold θ2 with non-target chromosomes is 0.3, that is, it is required that probe P i-1 has a similarity on non-target chromosomes less than or equal to 30%.

[0090] (1) The computer executes the steps: perform a second round of alignment and screening on the selected probes, and align the probes with the non-target chromosome library:

[0091] lastn - query probes.fasta - db nontarget_chromosome_db - out results_nontarget.txt

[0092] 4. Structure prediction and screening. For the probes that have passed the two rounds of alignment and screening, the stability of the probes also needs to be evaluated by the non-fold structure (non-folding structure with complicated secondary structure DNA domain) prediction tool NUPACK (http: / / www.nupack.org / ). For each probe P in the secondary probe screening set i-2 Evaluate the stability of probe P through the non-fold structure prediction tool i-2 to screen out probe P i-2 with a stability greater than or equal to the threshold θ3. The set of probes P i-3 obtained by screening is the stability probe screening set, where the stability probe screening conditions are as follows:

[0093] F(P i-2 ) ≥ θ3,

[0094] where F(P i-2 ) is the non-fold structure stability score of probe P i-2 ;

[0095] θ3 is the non-fold structure stability score threshold of probe P i-2 ; Set the non-fold structure stability score threshold θ3 to 0.7.

[0096] In addition, the number of repetitions of each probe P in the stability probe screening set i-3 in the centromere region and telomere region of the target chromosome and non-target chromosome is screened, and probe P i-3Probes with the number of repeats on the target chromosome greater than or equal to the number threshold r1 and the number of repeats on non-target chromosomes less than or equal to the number threshold r2 are screened to obtain the probe set P i-4 The set is the repeat number probe screening set, where the repeat number probe screening conditions are as follows:

[0097] R target (P i-3 ) ≥ r1 and R non-target (P i-3 ) ≤ r2,

[0098] In the formula, R target (P i-3 ) is the number of repeats of probe P i-3 in the centromeric region and telomeric region of the target chromosome, and R non-target (P i-3 ) is the number of repeats of probe P i-3 in the centromeric region and telomeric region of non-target chromosomes;

[0099] r1 is the repeat number threshold of probe P i-3 in the centromeric region and telomeric region of the target chromosome, and r2 is the repeat number threshold of probe P i-3 in the centromeric region and telomeric region of non-target chromosomes;

[0100] r1 is set to 5 times and r2 is set to 1 time, that is, the number of repeats of probe P i-3 in the centromere and telomere regions of the target chromosome is at least 5, while the number of repeats in the centromere and telomere regions of non-target chromosomes does not exceed 1 time.

[0101] (1) The computer executes the steps: According to the alignment results, use a Python script to perform non-fold structure screening and optimize probe performance:

[0102] python3 GENOME_FILTER.py --input results_target.txt --filter results_nontarget.txt.

[0103] 5. Final screening. Finally, the screened probes need to meet all of the following conditions:

[0104] ⅰ First-round alignment screening: S target (P i ) ≥ θ1 (set θ1 = 0.8);

[0105] ⅱ Second-round alignment screening: S non-target (P i-1 ) ≤ θ2 (set θ2 = 0.3);

[0106] ⅲ Non-fold structure prediction: F(P i-2 ) ≥ θ3 (set θ3 = 0.7);

[0107] ⅳ Repeat number screening: R target (P i-3 ) ≥ r1 and R non-target (P i-3 ) ≤ r2 (set r1 = 5, r2 = 1);

[0108] The probe P of the obtained repeat number probe screening set i-4 will be selected as the final qualified probe, and the probe P will be output i-4 , that is, a highly specific chromosomal probe sequence is obtained;

[0109] (1) The computer execution steps: After two rounds of alignment and screening, the first n bits (determine the number of probes according to actual needs, if the number of returned probes is small, all can be used) of the finally screened highly specific chromosomal probe sequence are output:

[0110] python3 FinalProbeSelector.py --input filtered_probes.fasta --output final_probes.fasta.

[0111] Example 2

[0112] A highly specific human chromosome 1 probe combination

[0113] The highly specific human chromosome 1 probe combination of this example is obtained by screening based on the screening method of Example 1.

[0114] The position of the centromeric region of human chromosome 1 on the chromosome is: Chromosome1: 122026460 - 125184587, the target chromosome is human chromosome 1, and the non-target chromosomes are the sum of the 23 pairs of chromosomes other than chromosome 1 in human chromosomes. The probe sequences finally screened by the screening method of Example 1 are shown in Table 1. The probe sequences are Chr1_1, Chr1_2, Chr1_3, Chr1_4, Chr1_5, Chr1_6, Chr1_7, Chr1_8, Chr1_9, and Chr1_10 respectively, and their nucleotide sequences are shown in SEQ ID NO: 1 - 10 respectively.

[0115] Table 1 Probe sequence combination

[0116]

[0117] Example 3

[0118] A kit

[0119] The kit of this embodiment contains the highly specific human chromosome 1 probe combination of Embodiment 2.

[0120] This kit can be used to detect human cell chromosomes.

[0121] Example 4

[0122] Specificity verification of the human chromosome 1 probe combination

[0123] Use the chromosome probe combination of Embodiment 2 or the kit of Embodiment 3 for specificity verification. Synthesize the probe sequences Chr1_1 to Chr1_10 and modify them with a fluorescent group (any one of Cy3, Cy3.5, Cy5, Cy5.5, Cy7, Cy7.5, Alexa555, Alexa546, and Alexa532, etc.), and mix them into mixprobe for hybridization verification on human cells.

[0124] The specific steps of hybridization are as follows:

[0125] 1) Boiling the slides: Prepare cell drops using human cells, and soak the prepared cell drops in 2×SSC buffer at 65±1°C for 30 min.

[0126] 2) Gradient dehydration: Soak the cell drops in ddH2O, 70% ethanol, 85% ethanol, and 100% ethanol solutions for 1 min each at room temperature.

[0127] 3) Hybridization (performed under light - proof conditions): After the cell drops are dry, add the pre - mixed and centrifuged probe to the target area, cover the coverslip avoiding air bubbles, seal the edge of the coverslip with mounting glue, and place it in a hybridization oven (a wet box can be used instead) for hybridization after the glue dries.

[0128] Hybridization procedure: Denature at 82°C for 10 min and hybridize at 45°C for 2 h.

[0129] 4) Washing (performed under light - proof conditions): After hybridization, place the cell drops in ddH2O, wait for the coverslip to fall off naturally, then place them in the pre - heated washing solution (0.3% NP40 / 0.4xSSC) and elute at 68°C for 2 - 4 min. Transfer the cell drops to ddH2O for washing for 1 min, transfer them to 100% anhydrous ethanol for soaking for 1 min, and then air - dry.

[0130] 5) Microscopic examination (performed under light - proof conditions): After sealing with DAPI, perform fluorescence microscopy to check the hybridization of the probe with the cells.

[0131] The hybridization results are as Figure 1As shown, normal human cells have two sets of chromosomes. Therefore, after hybridization, each cell has two fluorescent dots, and there are no obvious fluorescent dots in other regions. The results show that the human chromosome 1 probe combination of the present invention can specifically bind to the chromosomes of human cells and has high specificity and high sensitivity. The human chromosome 1 probe combination of the present invention can be used for basic research. According to the binding situation of the probe to the chromosome, it can be studied whether there are abnormalities in the chromosomes of human cells, and it can also be used for clinical disease detection.

[0132] Other parts not described in detail are all prior arts. Although the above embodiments have made a detailed description of the present invention, they are only a part of the embodiments of the present invention, rather than all embodiments. People can also obtain other embodiments based on these embodiments without creative efforts, and these embodiments all fall within the protection scope of the present invention.

Claims

1. A method for screening a combination of highly specific chromosome probes, characterized in that: The following steps are involved: (1) Extract the sequence from the centromere region and / or telomere region of the target chromosome to generate probe P i The collection is the probe set; (2) Each probe P in the probe set i Compare with the centromere region and / or telomere region of the target chromosome, select probes whose similarity with the target chromosome is greater than or equal to the threshold θ1, and select the probes P i-1 The set is the preliminary probe screening set; (3) Each probe P in the preliminary probe screening set i-1 Compare with non-target chromosomes and select probes whose similarity with non-target chromosomes is less than or equal to threshold θ2; the probes P obtained by screening i-2 The set is the secondary probe screening set; (4) Each probe P in the secondary probe screening set i-2 The stability of the probes was evaluated by using a non-fold structure prediction tool, and the probes with stability greater than or equal to the threshold θ3 were screened. i-3 The set is the stability probe screening set; (5) Each probe P in the stability probe screening set i-3 The number of repetitions in the centromere region and / or telomere region of the target chromosome and the non-target chromosome is screened, and the number of repetitions of the screening probes in the target chromosome is greater than or equal to the number threshold r1, and the number of repetitions in the non-target chromosome is less than or equal to the number threshold r2. The screened probe P i-4 The set is the probe screening set for the number of repetitions; (6) Finally, output the number of repetitions for each probe P in the probe screening set. i-4 , and obtain highly specific chromosome probe sequences.

2. The screening method according to claim 1, characterized in that: In step (1), each probe P in the probe set i The design formula is as follows: P i =S i [start:end]; Where P i is the probe, i = 1, 2, ..., n, S is the target chromosome sequence, [start, end] is the start and end position of the centromere region or telomeric region of the target chromosome, and n is the number of probes generated; In step (2), each probe P in the preliminary probe screening set i-1 The filter conditions are as follows: S target (P i )≥θ1, In the formula, S target (P i ) is the probe P i Similarity with the target chromosome, θ1 is the probe P i similarity threshold with the target chromosome; In step (3), each probe P in the secondary probe screening set i-2 The filter conditions are as follows: S non-target (P i-1 )≤θ2, In the formula, S non-target (P i-1 ) is the probe P i-1 Similarity with non-target chromosomes, θ2 is the probe P i-1 similarity threshold with non-target chromosomes; In step (4), each probe P in the stability probe screening set i-3 The filter conditions are as follows: F(P i-2 )≥θ3, In the formula, F(P i-2 ) is the probe P i-2 The non-fold structure stability score of θ3 is the probe P i-2 The stability score threshold of the non-fold structure; In step (5), each probe P in the probe screening set is repeated i-4 The filter conditions are as follows: R target (P i-3 )≥r1 and R non-target (P i-3 )≤r2, In the formula, R target (P i-3 ) is the probe P i-3 The number of repeats in the centromeric and / or telomeric regions of the target chromosome, R non-target (P i-3 ) is the probe P i-3 the number of repeats in the centromeric and / or telomeric regions of non-target chromosomes; r1 is probe P i-3 The repeat number threshold in the centromere region and / or telomeric region of the target chromosome, r2 is the probe P i-3 The repeat number threshold in the centromeric and / or telomeric regions of non-target chromosomes.

3. The screening method according to claim 1, characterized in that: The non-fold structure prediction tool is NUPACK; In step (2), the probe P i The similarity threshold θ1 with the target chromosome is 0.8; In step (3), probe P i-1 The similarity threshold θ2 with non-target chromosomes is 0.3; In step (4), probe P i-2 The stability score threshold θ3 of the non-fold structure is 0.7; In step (5), probe P i-3 The repeat number threshold r1 in the centromere region and / or telomeric region of the target chromosome is 5, and the probe P i-3 The repeat count threshold in the centromeric and / or telomeric regions of non-target chromosomes was 1.

4. Use of the screening method according to any one of claims 1 to 3 in screening chromosome probes with high specificity and high sensitivity.

5. A highly specific chromosome probe combination screened by the method according to any one of claims 1 to 3, characterized in that: The probe combination includes 10 probe sequences, namely Chr1_1, Chr1_2, Chr1_3, Chr1_4, Chr1_5, Chr1_6, Chr1_7, Chr1_8, Chr1_9 and Chr1_10, and their nucleotide sequences are shown in SEQ ID NO: 1 to SEQ ID NO: 10, respectively.

6. The probe combination according to claim 5, characterized in that: The chromosome is human chromosome 1.

7. The probe combination according to claim 6, characterized in that: The probe combination was screened from the centromere region of human chromosome 1.

8. Use of the probe combination according to any one of claims 5 to 7 in preparing a kit.

9. A kit, characterized in that: The kit comprises the probe combination according to claim 5.

10. Use of the kit according to claim 9 in detecting chromosomes of human cells.

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