BioReagent 级 ，适用于对基线干扰要求严格的色谱和分析工作流程。
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在色谱分析、有机合成和交叉偶联反应领域已被 0 篇同行评审文献引用。
λ 噬菌体是最早使用的克隆载体，其基因组是长度约为 50kb 的双链 DNA 分子，其在宿主细胞由两种生活途径：1、裂解生长：环状 DNA 分子在细胞内多次复制，合成大量噬菌体基因产物，装配成噬菌体颗粒，裂解宿主菌再进行下一次感染；2、溶源性生长：感染细胞内 λ 噬菌体 DNA 整合到宿主菌染色体 DNA 中与之一起复制，并遗传给子代细胞，宿主细胞不裂解。科研人员常常利用 λ 噬菌体裂解生长的特点，培养获取大量的 λ 噬菌体颗粒，并提取 λ 噬菌体 DNA。噬菌体载体广泛用于文库筛选，目的克隆培养获得大量的噬菌体颗粒需要提取 λ 噬菌体 DNA 来开展测序等后续工作，λ 噬菌体裂解培养物离心后的上清，首先用 RNaseA/DNaseⅠ 混合酶消化去除残留的宿主菌 DNA/RNA，沉淀收集噬菌体，通过酚异戊醇等提取，残留碎片通过沉淀离心去除，通过一系列快速的漂洗、离心的步骤，将盐、细胞代谢物、蛋白等杂质去除，获得 DNA 的量很高，但是纯度一般，但是足够用于大多数分子生物学实验。
λ 噬菌体基因组 DNA 提取试剂盒 (PEG 沉淀法) 是简便的 λ 噬菌体基因组 DNA 的试剂盒，其提取原理是通过 PEG 沉淀、酚异戊醇等提取，残留碎片通过沉淀离心去除，通过一系列快速的漂洗、离心步骤，将盐、细胞代谢物、蛋白等杂质去除，即可获得 λ 噬菌体基因组 DNA，可进行酶切、PCR 等下游实验。该试剂仅用于科研领域，不适用于临床诊断或其他用途。
组分
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自备材料
1、 离心管、低温离心机、摇床
2、 70%乙醇、氯仿或氯仿替代物
操作步骤(仅供参考)
以下操作以10ml噬菌体感染细菌培养上清液为例。
1、 准备工作：向RNase A和DNase I分别加入1ml的TE 缓冲液吹打，颠倒混匀，充分溶解RNase A和DNase I后，按照每次使用量分装－20℃冻存，6个月有效
2、 将氯仿滴加到λ噬菌体感染的液体培养液中(氯仿终浓度在0.1～0.5%)，37℃振荡培养5-10min。如观察到裂解发生，取12ml上述培养液转移至离心管，8000～10000g离心10min，去除细菌碎片，取上清液。一般建议8000g离心，如果效果不佳可以考虑10000g，但转速过高容易导致噬菌体也沉淀至管底，降低产量。
3、 取10ml上清液，加入5μl RNase A和10μl DNase I，充分混匀，37℃培养30min。
注意：噬菌体培养上清液会因生长和裂解情况不同致使残留的RNA/DNA量不液不同，RNase/DNase消化过度，可能减少产量；消化不完全，DNA、RNA可粘走部分噬菌体，一般RNase可以进行调整，可根据实际情况适当调节用量和消化时间。
4、 向上清液中加入5mL噬菌体沉淀液，摇匀至溶解，冰浴1h或4℃过夜。
5、 4℃ 10000g离心20min，弃上清液。
6、 加入1mlSM 缓冲液充分清洗管壁及沉淀，转移至新的离心管或微量离心管，加入40μl噬菌体裂解液，68℃培养15min。
7、 加入等体积蛋白清除液，轻轻混匀，12000g离心5min，取上清液。
8、 （备选步骤）转移上清液至新的离心管中，加入等体积预冷的噬菌体漂洗液，轻轻混匀，-20℃孵育1h，4℃ 12000g离心10min，弃上清液。
9、 加入适量的70%乙醇溶液，混匀，4℃ 8000g离心8min，弃上清液；如果有必要，可以重复1次该清洗步骤。
10、室温自然干燥DNA，加入适量 TE 缓冲液，-20℃保存；TE 缓冲液体积越大，DNA浓度越低。
注意事项
1、 用于裂解的噬菌体、宿主菌越新鲜，裂解越好、收获量越大。
2、 液体培养裂解时，到了时间裂解还没发生，可适当的提高温度或加大振摇速度。
3、 RNase/DNase消化过度，可能减少产量；消化不完全，可粘走部分噬菌体。
4、 噬菌体沉淀液在低温下易析出白色物质，可60℃以上温浴至完全溶解。
5、 噬菌体裂解液在低温下易结晶析出白色物质，可37℃温浴至完全溶解。
6、 试剂开封后请尽快使用，以防影响后续实验效果。
7、 为了您的安全和健康，请穿实验服并戴一次性手套操作。
有效期：12个月有效。低温运输，按要求保存。
Bacteriophage λ is one of the earliest cloning vectors. Its genome is a double‑stranded DNA molecule of approximately 50 kb. It exhibits two alternative life cycles inside host cells:
1. Lytic growth: The circular phage DNA replicates multiple times within the host cell, directing synthesis of abundant phage‑encoded gene products. These components assemble into mature phage particles, which lyse the host bacterium to initiate new rounds of infection.
2. Lysogenic growth: Upon infection, λ phage DNA integrates into the host chromosomal DNA, replicates together with the host genome, and is passed on to daughter cells without host cell lysis.
Researchers exploit the lytic‑growth property of bacteriophage λ to propagate large quantities of phage particles for subsequent genomic DNA isolation. Phage vectors are widely applied in library screening. Once target clones are cultured to generate sufficient phage particles, λ phage DNA is extracted for downstream applications such as DNA sequencing.
The supernatant harvested by centrifugation of λ‑phage lysate is first treated with a mixture of RNase A and DNase I to degrade residual host‑bacterial DNA and RNA. Phage particles are then collected by precipitation, followed by phenol‑isoamyl alcohol‑based extraction. Cellular debris is removed by centrifugal precipitation. A series of rapid washing and centrifugation steps eliminate contaminants including salts, cellular metabolites and proteins. This procedure yields high‑quantity DNA with moderate purity, which is adequate for most molecular‑biology experiments.
The Bacteriophage λ Genomic DNA Extraction Kit (PEG Precipitation Method) enables simple isolation of λ phage genomic DNA. Its underlying principle relies on PEG‑mediated precipitation, phenol‑isoamyl alcohol extraction, and removal of residual debris by centrifugation. Salts, cellular metabolites, proteins and other impurities are eliminated via sequential rapid washing and centrifugation, yielding λ phage genomic DNA suitable for downstream experiments including restriction digestion and PCR.
For research use only. Not intended for clinical diagnosis or other applications.
Component
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Reagents, consumables and Equipments not provided
1. Centrifuge tubes, refrigerated centrifuge, shaker
2. 70% ethanol, chloroform or chloroform substitute
Operating Procedures (For Reference Only)
The following protocol uses 10 mL supernatant of phage‑infected bacterial culture as an example.
1. Preparation: Add 1 mL TE Buffer to RNase A and DNase I respectively. Pipette up‑and‑down and invert to mix thoroughly to fully dissolve RNase A and DNase I. Aliquot according to the volume per usage, store at −20 °C. Valid for 6 months.
2. Add chloroform drop‑wise to liquid culture infected with λ phage to a final chloroform concentration of 0.1‑0.5%. Incubate with shaking at 37 °C for 5‑10 min. Once lysis is observed, transfer 12 mL of the culture into a centrifuge tube. Centrifuge at 8000‑10000 g for 10 min to remove bacterial debris and collect the supernatant. 8000 g is generally recommended. 10000 g may be used if results are unsatisfactory; however, excessive centrifugal force may co‑precipitate phage and reduce yield.
3. Take 10 mL supernatant, add 5 μL RNase A and 10 μL DNase I, mix well, and incubate at 37 °C for 30 min.
Note: Residual RNA/DNA content varies in phage culture supernatant depending on bacterial growth and lysis status. Over‑digestion by RNase/DNase may lower yield; incomplete digestion allows residual DNA and RNA to sequester part of the phage. RNase dosage can be adjusted; optimize dosage and digestion time according to actual experimental conditions.
4. Add 5 mL phage precipitation solution to the supernatant, mix until dissolved, and incubate on ice for 1 h or at 4 °C overnight.
5. Centrifuge at 10000 g for 20 min at 4 °C and discard the supernatant.
6. Add 1 mL SM Buffer to thoroughly rinse the tube wall and pellet. Transfer to a new centrifuge tube or microcentrifuge tube, add 40 μL phage lysis solution, and incubate at 68 °C for 15 min.
7. Add an equal volume of protein removal solution, mix gently, centrifuge at 12000 g for 5 min, and collect the supernatant.
8. (Optional step) Transfer supernatant to a new centrifuge tube, add an equal volume of pre‑chilled Phage Wash Buffer, mix gently, incubate at −20 °C for 1 h. Centrifuge at 12000 g for 10 min at 4 °C and discard supernatant.
9. Add an appropriate volume of 70% ethanol, mix, centrifuge at 8000 g for 8 min at 4 °C, discard supernatant. Repeat this washing step once if necessary.
10. Air‑dry DNA at room temperature. Resuspend in appropriate volume of TE Buffer and store at −20 °C. Larger volumes of TE Buffer result in lower DNA concentration.
Precautions
1. Fresher phage and host bacteria for lysis yield better lysis efficiency and higher harvest.
2. If lysis does not complete within the expected time in liquid culture, moderately increase incubation temperature or shaking speed.
3. Over‑digestion with RNase/DNase may reduce yield; incomplete digestion can sequester partial phage particles.
4. White precipitate may form in phage precipitation solution at low temperature. Warm the solution above 60 °C until fully dissolved.
5. White crystalline precipitate may form in phage lysis solution at low temperature. Warm at 37 °C until fully dissolved.
6. Use reagents promptly after opening to avoid compromising experimental performance.
7. For personal safety and health, wear a lab coat and disposable gloves during operation.
Shelf‑life
12 months. Transport under cold conditions; store as specified.2 months. Transport under cold conditions; store as specified.
| 储运单元货号 | 储运单元名称 | 组分货号 | 组分名称 | 物理外观 | 50T | 储存 |
| P1527075NT | λ噬菌体基因组DNA提取试剂盒 (PEG沉淀法)(常温组分) | P1527075A | 噬菌体裂解液 | 液体 | 2ml | Room temperature |
| P1527075B | 噬菌体漂洗液 | 液体 | 100ml | Room temperature | ||
| P1527075C | TE 缓冲液 | 液体 | 5ml | Room temperature | ||
| P1527075WT | λ噬菌体基因组DNA提取试剂盒 (PEG沉淀法)(2-8℃组分) | P1527075D | 噬菌体沉淀液 | 液体 | 250ml | 2-8℃. |
| P1527075E | SM 缓冲液 | 液体 | 50ml | 2-8℃. | ||
| P1527075F | 蛋白清除液 | 液体 | 100ml | 2-8℃.避光 | ||
| P1527075DT | λ噬菌体基因组DNA提取试剂盒 (PEG沉淀法)(-20℃组分) | P1527075G | RNase A | 固体 | 10mg | -20℃. |
| P1527075H | DNase I | 固体 | 10mg | -20℃. |
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