Ordering
| Item | Catalog # | Description | Quantity | Price (USD) | ||
|---|---|---|---|---|---|---|
| Plasmid | 100850 | Standard format: Plasmid sent in bacteria as agar stab | 1 | $75 | Add to Cart | |
| AAV1 | 100850-AAV1 | Virus (100 µL at titer ≥ 1×10¹³ vg/mL)and Plasmid.More Information | Add to Cart | |||
This material is available to academics and nonprofits only.
Backbone
- Vector backbonepAAV(Search Vector Database)
- Vector typeMammalian Expression, AAV
Growth in Bacteria
- Bacterial Resistance(s)Ampicillin
- Growth Temperature37°C
- Growth Strain(s)NEB Stable
- Copy numberUnknown
Gene/Insert
- Gene/Insert namejRCaMP1b
- Alt nameRCaMP1h variant 1519
- SpeciesSynthetic
- PromoterSynapsin
Cloning Information
- Cloning methodUnknown
- 5′ sequencing primerunknown (Common Sequencing Primers)
Resource Information
- Supplemental Documents
- pAAV.Syn.Flex.NES-jRCaMP1b.WPRE.SV40 (p3851)_Q.gb
- Terms and Licenses
- UBMTA
- genOway Notice of RIghts
- Industry Terms
- Not Available to Industry
Depositor Comments
This plasmid was previously available as pAAV.Syn.Flex.NES-jRCaMP1b.WPRE.SV40 (p3851) from the Penn Vector Core. This plasmid was created as part of the GENIE project at Janelia Research Campus.
Information for AAV1 (Catalog # 100850-AAV1)(Back to top)
Purpose
Ready-to-use AAV1 particles produced from pAAV.Syn.Flex.NES-jRCaMP1b.WPRE.SV40 (#100850). In addition to the viral particles, you will also receive purified pAAV.Syn.Flex.NES-jRCaMP1b.WPRE.SV40 plasmid DNA.
Syn-driven, Cre-dependent NES-jRCaMP1b calcium sensor. These AAV preparations are suitable purity for injection into animals.Delivery
- Volume100 µL
- Titer≥ 1×10¹³ vg/mL
- Pricing$350 USD for preparation of 100 µL virus + $30 USD for plasmid.
- StorageStore at -80℃. Thaw just before use and keep on ice.
- ShipmentViral particles are shipped frozen on dry ice. Plasmid DNA (≥ 200ng) will also be included in the shipment.
Viral Production & Use
- Packaging Plasmidsencode adenoviral helper sequences and AAV rep gene, AAV1 cap gene
- BufferPBS + 0.001% Pluronic F-68
- SerotypeAAV1
- PurificationIodixanol gradient ultracentrifugation
Biosafety
Requestor is responsible for compliance withtheir institution"s biosafety regulations.Lentivirus is generally considered BSL-2. AAV isgenerally considered BSL-1, but may requireBSL-2 handling depending on the insert.Biosafety Guide
Resource Information
- Terms and Licenses
- Ancillary Agreement for Penn Vectors
- Terms of Use for Viral Vectors
- Industry Terms
- Not Available to Industry
Viral Quality Control
- Addgene ensures high quality viral vectors by optimizing and standardizing production protocols and performing rigorous quality control (QC) (see a list of our QC assays). Thespecific QC assays performed varies for each viral lot. To learn which specific QC assays were performed on your lot, please contact us.
- Titer: the exact titer of your sample will be reported on the tube. The titer you see listed on this page is the guaranteed minimum titer. See how titers are measured.
Visit our viral production page for moreinformation.
Addgene Comments
Using FLEX vectors in vivo: LoxP sites in FLEX plasmids are known to recombine during DNA amplification and viral vector production, which may result in a minority of Cre-activated (i.e., "flipped") viral vectors. Addgene has measured this occurs in 0.01-0.03% of viral vectors in our typical production protocol. This can lead to a small number of cells exhibiting Cre-independent transgene expression in vivo. To address this, we recommend titrating to find the optimal AAV dosage required for Cre-dependent transgene expression and function in vivo. This may include reducing the viral vector dosage in order to reduce the likelihood of Cre-independent expression.
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实验需要请问有没有养THP-1细胞的同仁？能否寄一皿我，我在广州南方医科大学，不胜感激！
激光共聚焦的看活细胞皿哪里买呢
想问下各位大神，本人用皿养的PC-9GR细胞突然有大片飘起，也并没有成团状飘起，求解决方法？
如题~
已经基本养好了贴壁的原代细胞了，准备鉴定了，看到好多都在说细胞爬片的问题，我可以不做爬片直接在皿里操作吗？具体需要注意些什么呢？感谢各位大神
ps：大皿和6孔板里细胞密度、培养条件等等都几乎一样。
我看园子里有人提过类似的问题，但还没人给出答案，希望有经验或有想法的战友帮忙分析分析啊~谢谢！
同学说传代必须是一皿传多皿，一皿传一皿就不算传代~求高手指点~~
传代后5天，依然没有长满，怕影响活力，想传代和冻存，不知这样是否可以？？
做细胞实验快半年了，一直都还挺顺的，这次五一放完假回来，复苏一支HK2，操作都跟以前一样，没想到出了好多问题：
冻存方法：包裹棉花直接-80℃过夜，第二天转移到液氮
冻存时间：1个半月前冻存的细胞，密度保证没问题，冻存前状态也好
复苏方法：液氮取出后37℃水浴，约2分钟溶解，加入6倍体积的完全培养基，800转离心5分钟，弃去上清，1ml完全培养基重悬，转移入培养皿（进口一次性塑料培养大皿），补足完全培养基，培养箱培养
第二天看细胞全都没贴壁，但是也没死，聚集成团装飘着，没有污染。不想重新离心加重机械损伤，就一直试试看的心态放在培养箱里养着了。又重新复苏一支，还是一样的结果，全飘着没贴壁。不死心，就往前面复苏的那一皿里直接加了1ml的血清，相当于18%的血清比例，过了一天去看，这下细胞都贴壁了。没有另外添加血清的那一皿就还是没贴壁。
另外还有一支以前复苏的HK2，也是一样的方法复苏的，那次复苏很好，细胞基本没什么死的，也都贴壁了，养在皿里状态也不错，但是拿来铺板就还是不贴壁，同样的培基（10%血清），铺板就一个不贴，皿里就都可以贴上。
求助各位战友：
1、复苏不贴壁是为什么？
2、增加血清比例能使复苏的细胞贴壁，这样的细胞是不是可以认为状态并不好，以后的培养是不是要一直这么高比例的血清？还是可以培养一段时间逐步减少血清比例？
3、仍然是10%血清，为什么铺板就不贴壁，而皿里的就没事？是不是铺板的时候也要增加血清比例呢？那在板里干预的过程中是不是要一直保持高比例的血清培养？
4、我的冻存及复苏方法是否有错？我觉得我的HK2从形态、生长速度上来说状态应该是不错的，而且我已经更换了全新的培基、血清和双抗，重新配置了完全培养基，不知道为什么会出现这种不贴壁的问题
拜托各位集思广益，细胞实验已经为了这个不贴壁的问题停滞快2周，心急如焚啊，拜托各位！
