Sir, please help me confirm my understanding. What I understand is that through RFLP, we can see the relatedness between two samples by comparing the different lengths in two DNA fragments with the same probe binding sequence. The difference in length between two fragments is due to the presence or absence of specific restriction sites for certain restriction enzymes. Is this correct?
Restriction sites for almost all restriction enzymes are always there in genome. So any enzyme is always cleave at multiple sites of genome and produce tens to hundreds of fragments. You need not worry about what restriction site to target. Hope this helps.
@@XploreBio thank you but my guide has told me to use rflp only and I couldn't find snp in restriction site so please suggest how find restriction sites in my particular Dna sequence or how should I proceed
RFLP marker will restrict any DNA considering it has restriction site. If your aim is to see genetic variation in different samples, you use one or more RE. We expect that there would be SNP at some restriction sites and you would probably get variable fragments to be analyzed. If you need to discuss in detail, you may write to me at xplorebio@yahoo.com
sir can u exxplain how the probe sequence is designed i mean on what basis r they designed? what r they complementary to? the technique produce variable lenght fragments between rest enzymes cut sites.. this variable fragments sequence vary among individuial and will vary according to the no cut sites .. so what are the probes bind to exactly and in what seq... thanks in advance
Short, single- or low-copy genomic DNA or cDNA clones 500-2000bp are typically used as RFLP probes. Refer the following paper link: www.ncbi.nlm.nih.gov/probe/docs/techrflp/ For research purpose do refer to multiple papers.
If you are planning for genotyping work, I recommend you to use markers such as SSR, SNP. For genetic fidelity testing ISSR. Find the links below: 1. SSR marker: th-cam.com/video/iGN2tFCLPZ0/w-d-xo.html 2. Markers: th-cam.com/video/Quk-Dh65iHY/w-d-xo.html 3. ISSR: th-cam.com/video/3UahNA_4lBg/w-d-xo.html
RFLP uses an enzyme that breaks the DNA in specific points, so we get different lengths of fragments. Using electrophoresis these fragments can be separeted. Now it's the part where i get confused, the radioactive probes will bind to a very specif fragment of DNA, how can people be individualized if we are looking for the same fragments for everyone? If the probe binds to that specif sequence, shouldn't it be in the same place for every sample in the gel? Also, how is decided which probe to use?
You are right.Probes are specific. Also there are hundreds of restriction sites in genomic DNA. Difference in fragments will be there if there is mutation/variation in restriction site. This is possible if individuals are diverse.
Could you please explain this to me - if we get a skin cell at a crime scene and then we extract the dna from it, that's 6.6 billion base pairs right? Do we restrict the entire DNA (6.6 billion base pairs) with restriction enzymes and do an rflp analysis?
Thanks for asking the question. You are right, you need to extract DNA, restrict it, and detect it using labelled probes. You also need another sample to compare with.
thanks a lot.
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Thanks, that was a very concise and helpful video(:
Sir very helpful .thank u sir 😊
Thanks alot ..it was very nice nd easy to understand.😊
Thanks Amrita.
Sir, please help me confirm my understanding. What I understand is that through RFLP, we can see the relatedness between two samples by comparing the different lengths in two DNA fragments with the same probe binding sequence. The difference in length between two fragments is due to the presence or absence of specific restriction sites for certain restriction enzymes. Is this correct?
You got that right.
@@XploreBio thank you very much, Sir!
Very nice explanation 👌
Thanks..
Excellent
Very intelligible.
Thank you so much sir
All the best
1:12 a mutation in a TATA sequence: damn, individual 1 could be in serious gene-transcription problems ;)
Good..
Thank you
Please make videos on plant biotechnology
If we use this RFLP technique, how do we identity the restrictions site?
Restriction sites for almost all restriction enzymes are always there in genome. So any enzyme is always cleave at multiple sites of genome and produce tens to hundreds of fragments. You need not worry about what restriction site to target. Hope this helps.
Kindly distinguished the two method of conducting DNA fingerprinting
Please ask clearly.
Please make videos on seed technology
sir, please tell what to do if we do not have snp or difference within restriction site but we have use rflp only to detect that snp
RFLP will detect SNP only if its in the restriction site. Otherwise you need to use other methods to detect SNP.
@@XploreBio thank you but my guide has told me to use rflp only and I couldn't find snp in restriction site so please suggest how find restriction sites in my particular Dna sequence or how should I proceed
RFLP marker will restrict any DNA considering it has restriction site. If your aim is to see genetic variation in different samples, you use one or more RE. We expect that there would be SNP at some restriction sites and you would probably get variable fragments to be analyzed.
If you need to discuss in detail, you may write to me at xplorebio@yahoo.com
@@XploreBio thanks 😊
Please make videos on plant breeding
Thank you so much
sir can u exxplain how the probe sequence is designed i mean on what basis r they designed? what r they complementary to?
the technique produce variable lenght fragments between rest enzymes cut sites.. this variable fragments sequence vary among individuial and will vary according to the no cut sites .. so what are the probes bind to exactly and in what seq... thanks in advance
Short, single- or low-copy genomic DNA or cDNA clones 500-2000bp are typically used as RFLP probes.
Refer the following paper link:
www.ncbi.nlm.nih.gov/probe/docs/techrflp/
For research purpose do refer to multiple papers.
If you are planning for genotyping work, I recommend you to use markers such as SSR, SNP. For genetic fidelity testing ISSR. Find the links below:
1. SSR marker:
th-cam.com/video/iGN2tFCLPZ0/w-d-xo.html
2. Markers:
th-cam.com/video/Quk-Dh65iHY/w-d-xo.html
3. ISSR:
th-cam.com/video/3UahNA_4lBg/w-d-xo.html
RFLP uses an enzyme that breaks the DNA in specific points, so we get different lengths of fragments. Using electrophoresis these fragments can be separeted. Now it's the part where i get confused, the radioactive probes will bind to a very specif fragment of DNA, how can people be individualized if we are looking for the same fragments for everyone? If the probe binds to that specif sequence, shouldn't it be in the same place for every sample in the gel? Also, how is decided which probe to use?
You are right.Probes are specific. Also there are hundreds of restriction sites in genomic DNA. Difference in fragments will be there if there is mutation/variation in restriction site. This is possible if individuals are diverse.
Could you please explain this to me - if we get a skin cell at a crime scene and then we extract the dna from it, that's 6.6 billion base pairs right? Do we restrict the entire DNA (6.6 billion base pairs) with restriction enzymes and do an rflp analysis?
Thanks for asking the question. You are right, you need to extract DNA, restrict it, and detect it using labelled probes. You also need another sample to compare with.
And the human genome is 3.2 billion bp.
@@XploreBio 3.2+ on one strand & 3.2+ on the other no?
@@zoyeb100 thats why its base pairs
Thx man
Sir, do you have an email I need your help with a case study
XploreBio@yahoo.com
Your resolution is not good it is not clear......
In silico: th-cam.com/video/BkTRYMjyatA/w-d-xo.html
Thank you