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Early diagnosis and detection system used for BKV (BK polyomavirus), and application of early diagnosis and detection system

An early diagnosis and systematic technology, applied in the biological field, can solve problems such as not optimal target location, inability to effectively cover BKV strains, and poor sequence integrity, and achieve comprehensive and reliable detection results, high sensitivity, and wide coverage

Pending Publication Date: 2020-01-14
上海公卫生物医药有限公司
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  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

[0004] However, the existing systems all have the following defects: 1. The target position is not optimal
By comparing and arranging the 315 pieces of NCBI so far (one of which has poor sequence integrity and is not included in the sequence array below), plus the 12 BKV whole genome sequences we sequenced, and then brought in the reported primers and The probe sequences were compared, and we found that these primers and probes have been unable to effectively cover all BKV strains, and the primer probe sequences (GC content and annealing temperature, etc.) of some systems also need to be optimized (Table 2)

Method used

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  • Early diagnosis and detection system used for BKV (BK polyomavirus), and application of early diagnosis and detection system
  • Early diagnosis and detection system used for BKV (BK polyomavirus), and application of early diagnosis and detection system
  • Early diagnosis and detection system used for BKV (BK polyomavirus), and application of early diagnosis and detection system

Examples

Experimental program
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Embodiment 1

[0043] Example 1 Designs the primers and probe sequences of BKV

[0044] (1) Align existing BKV sequences and find out 4 highly conserved regions of BKV sequences.

[0045] Selecting highly conserved sequences from the BKV genome is the primary task of designing specific primers and probes. The genome of BKV is divided into early coding region (including LT and sT genes), late coding region (including Agno, VP1, VP2 and VP3 genes) and non-coding regulatory region (NCCR) ( figure 2 A). Since point mutations, deletions and recombinations often occur in non-coding regulatory regions, the difficulty of sequence alignment is increased. Therefore, we edited all the existing 327 BKV sequences and removed the NCCR sequence, that is, we arranged from the start codon of the Agno gene at the 5' end of the late coding region to the 5' end of the early coding region (early and late coding region The sequence direction is reversed, so up to the start codons of sT and LT here). We compa...

Embodiment 2

[0066] Example 2 checks the primer and probe sequences of BKV

[0067] (1) Four groups of qPCR primers and probes were tested using the urine and serum of kidney transplant recipients.

[0068] We selected urine and serum samples from 6 kidney transplant recipients, extracted DNA, and performed quantitative PCR detection using the above-mentioned self-designed 4-group system. The results showed that the detection level and stability of VP2 and VP2 / 3 were relatively good ( image 3 ). In particular, the probe region of VP2 / 3 is a highly specific sequence of BKV, which completely avoids the interference of JCV during detection, so it is very suitable as a new system for BKV fluorescent quantitative PCR detection.

[0069] image 3 .VP2 / 3 system can be used as a new system for BKV fluorescent quantitative PCR detection. The samples in the picture are from 6 kidney transplant recipients. Among them, S stands for serum (Serum), and U stands for urine (Urine). In sample No. 3, se...

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Abstract

The invention discloses an early diagnosis and detection system used for a BKV (BK polyomavirus), and application of the early diagnosis and detection system, and particularly provides a primer probesystem used for the early diagnosis and detection of the BKV, a kit including the above primer probe, and the purposes of the primer probe system and the kit. The primer probe system disclosed by theinvention has a wider coverage degree, high specificity, higher sensitivity and a larger clinic application potential, and is favorable for effectively preventing BKV related diseases, and the qualityof life of renal transplant recipients can be improved.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to an early diagnosis and detection system for BKV and its application. Background technique [0002] BK polyomavirus (BK polyomavirus, BKV) is an opportunistic virus of latent infection, and the infection rate in the general population is as high as 85%. Once BKV is activated, it will seriously threaten immunocompromised people (immunosuppressant users after organ transplantation, HIV-infected patients, pregnant women and the elderly). Including: 1) It can cause BKV-related nephropathy in 1-10% of kidney transplant recipients, and if the activation of BKV is not inhibited in time and effectively, it will lead to transplanted kidney failure; 2) It can cause bleeding in about 10% of hematopoietic stem cell transplant recipients 3) BKV infection increases the risk of some urinary system tumors (such as bladder cancer). At present, there is no effective anti-BKV means except reducing imm...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/6851C12N15/11C12R1/93
CPCC12Q1/701C12Q1/6851C12Q2531/113C12Q2563/107C12N15/11C12Q1/70C12Q1/68Y02A50/30
Inventor 朱同玉吴楠楠石业静戎瑞明许明代发辉梅雪高文君曾毅刚唐群业胡春兰
Owner 上海公卫生物医药有限公司
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