To your original question, Ibrahim: The "lysis+binding buffer" of Quiagen Kits contain high concentrated chaotropic salts (if you are brave, you can spread a tiny amount on your finger and tast it). GATC or Gu-HCl disrupt any hydrophilic bonds, therefore lysing the cell membrane, linearizing nucleic acids but at the same time completely desatbilizing RNAses. Therefore, while in the lysis buffer, RNA is perfectly protected.

The high salt concentration is also essential for binding of RNA/DNA to the positively charged columns (virtually a sort of porous glass). If you finally flush the column with low-ionic strength buffer (or water), the nucleic acids are released again and can be eluted.

The only limitations for the kit that I am aware is the size of the RNA species and potential overloading. It is always better to start with lower amounts of tissue or cells, in particular if the lysate contains a lot of protein, proteoglycans, lipids and so on.

RNA shorter than about 100 nt bind poorly to the column, therefore they are lost or outcompeted by the longer RNAs.

Michael

Hi, from which Quiagen Kit you have this buffer ?

Most lysis buffer for cells as part of DNA/RNA-extraction kits contain chaotropic salts like GATC or guanidium HCl or sodium-iodide.

You can see this pdf u can got ur answer

https://repository.library.georgetown.edu/bitstream/.../leshinJonathan.pdf?...

Dear Siamak, the link does not work in my hands.

Dear Vivian i can sed you the pdf file. But i dont know how can send from researchgate?

The paper you are referring is here:

https://repository.library.georgetown.edu/bitstream/handle/10822/553187/leshinJonathan.pdf?sequence=1 (hope ResearchGate shows the proper link, otherwise seach Google for "library georgetown leshinJonathan.pdf"

But it wont be of any help, since RLP in this text stands for Ricin-Like Protein, not for a Quiagen Buffer.

Are you sure you are talking about RLP, and not ALP buffer ? Since none of the Quiagen technical ressources contain any information on RLP.

Getting back to the purpose of the buffer itself, this proprietary lysis buffer is similar to MoBIO and others' lysis buffer in that it contains detergent, perhaps SDS or a Triton component, with an osmoprotectant (glucose) in a Tris buffered solution. I have a specific recipe for something similar and can pass along if you need to make more of this solution for your kit.

To your original question, Ibrahim: The "lysis+binding buffer" of Quiagen Kits contain high concentrated chaotropic salts (if you are brave, you can spread a tiny amount on your finger and tast it). GATC or Gu-HCl disrupt any hydrophilic bonds, therefore lysing the cell membrane, linearizing nucleic acids but at the same time completely desatbilizing RNAses. Therefore, while in the lysis buffer, RNA is perfectly protected.

The high salt concentration is also essential for binding of RNA/DNA to the positively charged columns (virtually a sort of porous glass). If you finally flush the column with low-ionic strength buffer (or water), the nucleic acids are released again and can be eluted.

The only limitations for the kit that I am aware is the size of the RNA species and potential overloading. It is always better to start with lower amounts of tissue or cells, in particular if the lysate contains a lot of protein, proteoglycans, lipids and so on.

RNA shorter than about 100 nt bind poorly to the column, therefore they are lost or outcompeted by the longer RNAs.

Michael

Michael Rosemann provided the full answer to Ibrahim's question. I would only add, that the RLT Buffer is one of the buffers from RNeasy kit which is used, as Michael has mentioned, for isolating total RNA from different types of cells and tissues. The RNeasy handbook states about RLT Buffer: "Buffer RLT contains guanidine thiocyanate". You can add beta-mercaptoethanol to the RLT to inhibit the RNases.

Are you mean RLT buffer?The exact composition of Buffer RLT is confidential. This buffer is a proprietary component of RNeasy Kits. Buffer RLT contains a high concentration of guanidine isothiocycanate, which supports the binding of RNA to the silica membrane. Buffer RLT can be purchased separately (cat. no. 79216)

Thanks A lot Michael Rosemann, I think your answer is a good one,

Yes I mean RLP of Qiagen kits for total RNA extraction from blood,

Thanks everybody who tried to help through her/ his answer

Dear Ibrahim

Lysis buffers mostly contains guanidine thiocyyanate (5M-10M) which is dissolved in Tris-Hcl (0.1 M) + 0.5M EDTA + Tween-20 or Triton X-100. As Michael said it would disrupt any hydrophilic bonds, therefore lysing the cell membrane, linearizing nucleic acids but at the same time completely desatbilizing RNAses.

I have a relative question about buffer RLT. I am using Qiagen Allprep kit to isolate RNA and DNA from brain white matter. Since this is a lipid rich tissue there is a white fatty layer after homogenization with buffer RLT. I am getting decent DNA yield but no RNA. My guess is this is because of the fatty layer interference. I try to remove with pipet but not easy to get all of it out. How can I get good RNA yield?

can i use sds as a alternative of ATL in qiagen blood DNA mini kit in initial step. can anyone tell me? i have no ATL buffer.

Sukrutha C,

Have you ever tried QIAzol from QIAGEN prior performing RNA isolation? In the product details in their site is written that QIAzol can be used to lyse all classes of tissues but is optimized for lysis of fatty tissues, such as brain and adipose tissues. Or try to homogenize with TRIzol, that have the same principle and can working as well in brain tissues.

Take a look at the link and see if that product satisfy your necessities.

https://www.qiagen.com/no/shop/sample-technologies/rna/qiazol-lysis-reagent/#productdetails