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Antisense nucleic acid of resistant and tolerant dimethoxyphenecillin staphylococcus aureus drug resistant gene

A methicillin-resistant Staphylococcus technology, which is applied in the field of antisense nucleic acid against methicillin-resistant Staphylococcus aureus drug-resistant genes, can solve the problem of not being a good target site, and achieve the suppression of MRSA drug resistance. The effect of applying the foreground

Inactive Publication Date: 2011-01-12
FOURTH MILITARY MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Usually the lower their negative value, the more stable, not a good target site

Method used

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  • Antisense nucleic acid of resistant and tolerant dimethoxyphenecillin staphylococcus aureus drug resistant gene
  • Antisense nucleic acid of resistant and tolerant dimethoxyphenecillin staphylococcus aureus drug resistant gene
  • Antisense nucleic acid of resistant and tolerant dimethoxyphenecillin staphylococcus aureus drug resistant gene

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Example 1 Design of antisense thio-oligodeoxynucleotides (PS-ODNs) and thio-deoxyribozymes (PS-DRz) against mecR1 or blaR1 mRNA:

[0029] The sequence of the MRSA drug-resistant gene mecR1 / mecA (gene number gi:2791983) was retrieved from GENBANK. The sequence is 9047bP in full length and encodes 8 open reading frames, including 3 mec regions and 5 ORF regions. The sequence between 1615bp and 3372bp is the mecR1 gene sequence, and the sequence between 3472bp and 5478bp is the mecA gene sequence. The full length of mecR1 is 1758bp, which is the upstream gene of mecA, which can encode MecR1 protein and regulate the expression of mecA; the full length of mecA is 2007bp, which encodes the penicillin binding protein PBP2a. The genome sequence of blaR1 / blaZ (gene number gi:33390917) was retrieved by the same method. The sequence is 3080 bp in full length and encodes three open reading frames, namely blaZ, blaR1 and blal3 regions. The sequence between 953bp and 2710bp is the b...

Embodiment 2

[0034] Design and synthesis of embodiment 2PCR primers:

[0035] According to the gene sequences of mecR1, mecA, blaR1, blaZ and 16srRNA of MRSA reported in GeneBank, five pairs of specific amplification primers for mecR1, mecA, blaR1, blaZ and 16srRNA were designed with the software PrimerPremier5.0. The primers were verified to be highly specific by BLAST software, and were synthesized by Shanghai Bioengineering Technology Service Co., Ltd. The sequences are shown in Table 2.

[0036] Table 2 designed and synthesized amplification primers

[0037]

Embodiment 3

[0038] Example 3 Preparation of electrocompetent WHO-2:

[0039] The methicillin-resistant Staphylococcus aureus strain WHO-2 was provided by the China Antimicrobial Resistance Surveillance Center. Transplant 1ml of freshly cultured overnight WHO-2 into 100ml of nutrient broth medium, culture at 37°C and 210rpm until mid-logarithmic growth, and measure its OD 600 =0.55~0.6. After 20 minutes in ice bath, subpackage (40ml / centrifuge tube), centrifuge at 6000rpm for 10min, discard the supernatant, mix and resuspend the bacterial pellet with 40ml ice-bathed deionized water, centrifuge at 6000rpm for 10min, discard the supernatant, repeat 2 times. Afterwards, the bacterial pellet was mixed and resuspended with 20 ml ice-bathed 10% glycerol, centrifuged at 6000 rpm for 10 min, and the supernatant was discarded. The bacterial pellet was mixed and resuspended with 1 ml of ice-bathed 10% glycerol, centrifuged at 6000 rpm for 10 min, the supernatant was discarded, and repeated twice....

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Abstract

The invention relates to antisense oligonucleotides aiming to methicillin-resistant Staphylococcus aureus (MRSA) resistance gene mecR1 / mecA and blaR1 / blaZ and a preparation of the drugs with the antisense oligonucleotides and the application thereof. The antisense oligonucleotides comprise antisense phosphorothiate oligodeoxynucleotide (PS-ODNs) and thiodeoxyribozyme (PS-DRz). The invention is characterized in that the combination of the specificity and the various areas of MRSA resistance gene mecR1 / mecA and blaR1 / blaZ blocks the expression of the resistance gene, the base sequence of the antisense nucleic acid is PS-ODNs01-40, the antisense nucleic acid can combine with the specific site of the target gene, thereby inhibiting the expression of the resistance gene, causing MRSA to restore the sensitivity of the lactam antibiotics and achieving the purpose of effective fighting with the drug resistance of MRSA.

Description

technical field [0001] The invention relates to a class of antisense nucleic acid sequences for drug-resistant genes mecR1 / mecA and blaR1 / blaZ of methicillin-resistant Staphylococcus aureus (MRSA), preparation of medicines containing the antisense nucleic acid sequences and use thereof. Background technique [0002] At present, the problem of bacterial drug resistance has intensified around the world, and many bacterial infectious diseases that were once curable are becoming incurable diseases. MRSA is an important pathogenic bacterium for nosocomial infection and community infection. Since MRSA was first discovered in 1961, especially after the 1980s, the infection rate of MRSA has shown a clear upward trend in most areas. For example, in the United States, the infection rate of MRSA increased from 1974 to 2% in 2003 to nearly 60% in 2003, an increase of almost 30 times in 30 years; Italy rose from 6% in 1981 to 26% in 1996. With the continuous improvement of MRSA infectio...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07H21/04C12N15/11A61K48/00C12N15/113
Inventor 罗晓星孟静茹侯征扈本荃王慧贾敏
Owner FOURTH MILITARY MEDICAL UNIVERSITY
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