BioVector® Xen29 甲氧西林敏感性金黄色葡萄球菌活体成像表达菌株 / Staphylococcus aureus Xen29 Strain

一、 背景与来源说明 / Background & Origin

Staphylococcus aureus Xen29（常用名 Xen29）是一种经基因工程改造的甲氧西林敏感性金黄色葡萄球菌（MSSA）生物发光报道菌株。该菌株衍生自亲本株 ATCC 12600，通过在其染色体中稳定整合发光发光杆菌（Photorhabdus luminescens）的完整发光基因盒 luxABCDE 表达构建而成。

由于 luxABCDE 操纵子同时包含产生发光酶（Luciferase）及合成其底物（脂醛）所需的全套基因，Xen29 在代谢活性良好的状态下无需外源添加荧光素底物即可持续稳定发出绿色/黄绿色生物发光（峰值约 490 nm）。该菌株被广泛应用于小动物活体成像系统（IVIS® Imaging System），开展金黄色葡萄球菌感染模型构建、体体内外抗生素药效评估及生物膜（Biofilm）形成动态监测。

二、 菌株生物学特征 / Biological Characteristics

• 物种分类 / Taxonomy：金黄色葡萄球菌 (Staphylococcus aureus)

• 生物安全等级 / Biosafety Level：BSL-2（致病性革兰氏阳性球菌，需在 BSL-2 实验室操作）

• 亲本来源 / Parental Strain：ATCC 12600 (MSSA)

• 抗生素敏感性 / Antibiogram：

  • 甲氧西林敏感（MSSA，对苯唑西林 Oxacillin、氨苄西林等敏感）

  • 筛选抗性：染色体整合位点或辅助筛选包含卡那霉素 (Kanamycin, 200 ug/mL) 标记，可用于维持培养防杂菌污染

• 核心发光元件 / Luminescence System：染色体整合 Photorhabdus luminescensluxABCDE operon（自主发光，无须依赖外源 Luc Substrate）

• 主要应用 / Key Applications：

  • IVIS 活体成像：小鼠软组织感染、败血症、灼伤创面感染及骨髓炎模型

  • 生物膜研究：植入物（如导管、植入泵）表面生物膜消长动态检测

  • 药物筛选：高通量体外/体内抗葡萄球菌新药与抗生素实时药效评估

三、 培养条件与保存指南 / Culture & Handling Protocols

1. 常用培养基 / Culture Media

• 液体培养基：TSB (Tryptic Soy Broth) 或 LB 培养基，通常添加 200 ug/mL 卡那霉素 (Kanamycin)

• 固体平板：TSA (Tryptic Soy Agar) 或 5% 羊血琼脂平板 (Blood Agar) + 200 ug/mL Kanamycin

• 培养条件：37 摄氏度，好氧培养，液体培养时 200–240 rpm 震荡

2. 复苏与表达检测 / Recovery & Bioluminescence Detection

1. 从 -80 摄氏度冻存柜中取出 Xen29 冻存管，置于冰上。

2. 用无菌接种环划线接种于含 200 ug/mL Kanamycin 的 TSA 或血琼脂平板上。

3. 37 摄氏度倒置培养 16–18 小时，长出圆形、表面光滑且带溶血环的单菌落。

4. 挑取单菌落接种至含卡那霉素的 TSB 中，37 摄氏度震荡培养至对数生长期（OD600 约 0.5–0.8）。

5. 发光验证：将菌液置于暗室或 IVIS 系统中，可直接检测到明亮的自发荧光信号（Signal Intensity 与活菌 CFU 呈良好线性相关）。

3. 菌种冻存保存 / Cryopreservation

• 取对数生长期菌液，加入无菌甘油至终浓度 15% 至 20%。

• 混匀后分装于无菌冻存管，直接置于 -80 摄氏度 超低温冰箱长期保存。

四、 技术指标简表 / Technical Data Summary

BioVector® Staphylococcus aureus Xen29 Strain

1. Background and Origin

Staphylococcus aureus Xen29 is a genetically engineered methicillin-susceptible Staphylococcus aureus (MSSA) bioluminescent reporter strain. Derived from the parental strain ATCC 12600, Xen29 carries a stably integrated copy of the full Photorhabdus luminescensluxABCDE operon on its bacterial chromosome.

Because the luxABCDE cassette encodes both the luciferase enzyme and the enzymes required for fatty aldehyde substrate synthesis, active metabolizing Xen29 cells emit a continuous, stable bioluminescent signal (peak wavelength ~490 nm) without requiring exogenously added substrates.Xen29 is widely utilized with IVIS® imaging systems to monitor live bacterial load, soft tissue/device-related infection dynamics, and antimicrobial efficacy in real time.

2. Biological Characteristics

• Taxonomy: Staphylococcus aureus

• Biosafety Level: BSL-2 (requires containment)

• Parental Origin: ATCC 12600 (MSSA)

• Antimicrobial Susceptibility:

  • Methicillin-sensitive (MSSA; susceptible to oxacillin, ampicillin, etc.)

  • Selective Marker: Kanamycin resistance (200 ug/mL Kanamycin) for reporter selection and contamination prevention

• Bioluminescent System: Integrated Photorhabdus luminescensluxABCDE operon (autonomous bioluminescence)

• Applications:

  • In Vivo IVIS Imaging: Murine soft tissue, scald wound, sepsis, and device implant infection models

  • Biofilm Analysis: Non-invasive tracking of bacterial biofilm formation on catheter/implant surfaces

  • Drug Discovery: High-throughput in vitro screening and real-time in vivo antimicrobial drug efficacy monitoring

3. Culture and Handling Protocols

Growth Media

• Liquid Broth: Tryptic Soy Broth (TSB) or LB Broth supplemented with 200 ug/mL Kanamycin

• Solid Media: Tryptic Soy Agar (TSA) or 5 percent Sheep Blood Agar supplemented with 200 ug/mL Kanamycin

• Incubation Conditions: Aerobic at 37 degrees Celsius with agitation at 200–240 rpm for broth cultures

Reconstitution & Bioluminescence Assay

1. Streak a small freeze-thaw portion from the -80 degrees Celsius stock onto selective agar plates (TSA + Kanamycin).

2. Incubate at 37 degrees Celsius for 16 to 18 hours until single colonies appear.

3. Pick a single colony into fresh TSB broth containing 200 ug/mL Kanamycin and grow to log phase (OD600 ~0.5 to 0.8).

4. Signal Verification: Photons can be directly captured using an IVIS Spectrum camera or luminometer without adding exogenous luciferin.

Cryopreservation

• Mix log-phase bacterial suspension with sterile glycerol to a final concentration of 15 to 20 percent.

• Aliquot into cryovials and store immediately at -80 degrees Celsius.

4. Technical Data Summary

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