GENETIC TEACHER

Microbial Molecular Ecology Investigate molecular interaction and molecular functions of microorganisms and virus within diverse environments and hosts, Microbial Diversity and Community Structure Investigate microbial community diversity and composition in various environments (Soil and Water) to identify and classify microorganisms, Environmental and Applied Microbiology study microbial process in natural and artificial environments to develop and assess microorganisms in environmental sustainability, Improvement Molecular Tools use bioinformatics analysis to analyze large scale of microbial and viral multi-omics data (Watch Related Video in #geneticteacher) #geneticteacher

Amplified Fragment Length Polymorphism (AFLP Marker) use restriction enzymes to digest genomic DNA and adaptors to ligate digested DNA fragments and pre-selective and selective PCR amplification of ligated fragments and vertical gel electrophoresis by using denature polyacrylamide gel to separate amplified fragments, DNA digest with two restriction enzymes which cut un-evenly overhang sticky ends such as., Mse-1 (Cut Frequently) and ECOR-1 (Cut Rarely), Component of restriction digestion genomic DNA in microliter: Genomic DNA and any kind of arbitrary primer (OPA) and Mse-1 and BSA (Come with Mse-1) and ECOR-1 and deionized water then vortex samples and incubate at 37 degree Celsius for 3 hours with agitate every hour after that inactivate enzymes at 70 degree Celsius for 15 minutes Next use adaptors of known sequence as priming sites to ligate sticky ends of digested DNA fragments, Ligation Mix Adaptors of AFLP Marker: ECOR-1 adaptor and Mse-1 adaptor and T4 DNA ligase 10X buffer and...

AFLP Marker detect genomic restriction fragments using PCR amplification for detection restriction fragments to display presence (1) or absence (0) restricted fragments while RFLP Marker detect length of restricted fragments using southern blot hybridization to detect restricted fragments to display length differences of restricted fragments (Watch Related Video in #geneticteacher) #geneticteacher

AFLP Marker based on selective PCR amplification of digested DNA fragments after digest DNA and ligate to synthetic adaptors and amplifying by primers that carry selective nucleotides at their 3 ends to detect polymorphism by changes in restriction sites or their neighboring regions (Watch Related Video in #geneticteacher) #geneticteacher

AFLP Marker use in genetic diversity studies and germplasm collection studies and genome mapping and gene tagging (Watch Related Video in #geneticteacher) #geneticteacher

AFLP Marker is a dominant inheritance marker because it doesn't distinguish between heterozygous and homozygous alleles and generate huge information leading to require technically skills in laboratory and data analysis so it need more time therefore it consider complex and costly marker (Watch Related Video in #geneticteacher) #geneticteacher

AFLP Marker allow quick scan of whole genome because large number of generated bands give highly informative fingerprint and it doesn't require prior sequence information or probe therefore AFLP Marker classify as highly polymorphism and reproducible hence it consider extremely useful in creating quick genetic map and transcript profile (Watch Related Video in #geneticteacher) #geneticteacher

Polymorphism of AFLP Marker detect by vertical gel electrophoresis using denature polyacrylamide gel to visualize separated fragments by radioisotopes or fluorescent dyes or silver stain to score bands as presence (1) or absence (0) (Watch Related Video in #geneticteacher) #geneticteacher

Component of selective PCR amplification: Diluted DNA template from pre-selective PCR reaction and ECOR-1 with selective nucleotide (AAC) and Mse-1 with selective nucleotide (AAC) and dNTPs and 10X PCR buffer and MgCl2 and Taq DNA polymerase and deionized water, Selective PCR reaction include two phases (First phase involve 12 Cycles with decrease temperature by 0.7 degree Celsius) while (Second phase include 23 Cycles) and after finish all phases add 8 microliters Formamide loading buffer to PCR product (Watch Related Video in #geneticteacher) #geneticteacher

Component of pre-selective PCR amplification: DNA template from restriction/ligation and ECOR-1 adaptor with Adenosine nucleotide (As primer) and Mse-1 adaptor with Cytosine nucleotide (As primer) and dNTPs and 10X PCR buffer and MgCl2 and Taq DNA polymerase and deionized water, Reaction of Pre-selective PCR amplification (26 Cycles) and after finish add 100 microliter of sterile water to PCR product, After that undergo selective PCR amplification by adding two nucleotide of Adenosine and one nucleotide of Cytosine to primer at 3 ends to again select subset fragments (Be Attention) More selective nucleotide detect less polymorphism so must only add three nucleotide (Watch Related Video in #geneticteacher) #geneticteacher

Component of ECOR-1 adaptor and Mse-1 adaptor and PCR profile for each mix reagent: One selective PCR consist of adaptor sequences of each restriction enzymes and one arbitrary nucleotide (OPA) to amplify only subset of digested DNA fragments and after finish store tubes at minus 20 degree Celsius after that perform pre-selective PCR amplification to amplify specific subsets of digested DNA fragments using combination of selective primers at 3 ends which are Adenosine nucleotide to ECOR-1 adaptor and Cytosine nucleotide to Mse-1 adaptor that complementary to adaptors and restriction sites to select only subset fragments (Watch Related Video in #geneticteacher) #geneticteacher

Ligation Mix Adaptors of AFLP Marker: ECOR-1 adaptor and Mse-1 adaptor and T4 DNA ligase 10X buffer and T4 DNA ligase units and deionized water add 10 microliters of ligation mix to 50 microliter digested DNA after that vortex samples and incubate at room temperature for 3 hours with agitate every hour (Watch Related Video in #geneticteacher) #geneticteacher

DNA digest with two restriction enzymes which cut un-evenly overhang sticky ends such as., Mse-1 (Cut Frequently) and ECOR-1 (Cut Rarely), Component of restriction digestion genomic DNA in microliter: Genomic DNA and any kind of arbitrary primer (OPA) and Mse-1 and BSA (Come with Mse-1) and ECOR-1 and deionized water then vortex samples and incubate at 37 degree Celsius for 3 hours with agitate every hour after that inactivate enzymes at 70 degree Celsius for 15 minutes Next use adaptors of known sequence as priming sites to ligate sticky ends of digested DNA fragments (Watch Related Video in #geneticteacher) #geneticteacher

Amplified Fragment Length Polymorphism (AFLP marker) use restriction enzymes to digest genomic DNA and adaptors to ligate digested DNA fragments and pre-selective and selective PCR amplification of ligated fragments and vertical gel electrophoresis by using denature polyacrylamide gel to separate amplified fragments (Watch Related Video in #geneticteacher) #geneticteacher

Random Amplified Polymorphic DNA (RAPD marker) Amplify genomic DNA randomly between two identical short sequence with single short oligonucleotide primer (10 base pair) based on PCR amplification by using low annealing temperature (35 to 45 degree Celsius) to increase sequence amplification without exactly complementary to primer sequence therefore changes in DNA sequence at each locus in RAPD marker assess by presence (1) or absence (0) bands on gel Hence RAPD polymorphism occur due to base substitution at primer binding sites or indels in regions between binding sites and the mismatch between primer and DNA template results decrease amount of PCR product, Initial denaturation start at 94 degree Celsius for 10 minutes and denaturation temperature should be at 94 degree Celsius for 1 minute and annealing temperature should be at 37 to 45 degree Celsius for1 minute and extension temperature should be at 72 degree Celsius for 1 minute and final extension should be at 72 degree Celsiu...

Random Amplified Polymorphic DNA (RAPD marker) Amplify genomic DNA randomly between two identical short sequence with single short oligonucleotide primer (10 base pair) based on PCR amplification by using low annealing temperature (35 to 45 degree Celsius) to increase sequence amplification without exactly complementary to primer sequence therefore changes in DNA sequence at each locus in RAPD marker assess by presence (1) or absence (0) bands on gel Hence RAPD polymorphism occur due to base substitution at primer binding sites or indels in regions between binding sites and the mismatch between primer and DNA template results decrease amount of PCR product (Watch Related Video in #geneticteacher) #geneticteacher

RAPD marker is a PCR based technique in which single short oligo-nucleotide primer use to amplify random sequences from DNA template and it's product separate by horizontal electrophoresis and visualize by ultraviolet illuminate of ethidium bromide stained agarose gel so polymorphism of amplified sequence corresponds to genetic differences (Watch Related Video in #geneticteacher) #geneticteacher

RAPD marker use in genetic diversity studies and germplasm characterization and genetic structure of population and genome mapping (Watch Related Video in #geneticteacher) #geneticteacher

RAPD marker classify as a dominant marker inheritance because homozygous and heterozygous states can not be differentiate on gel and not locus specific because short primer can amplify any of random sequence in genome and it's bands are very sensitive to reaction conditions and DNA quality and PCR temperature so RAPD marker Poor Reproducibly and sensitive (Watch Related Video in #geneticteacher) #geneticteacher

RAPD marker provide quick screen of DNA sequence without require any previous sequence information by using small amount of DNA to get high polymorphism therefore RAPD marker classify as simple and rapid and cheap marker (Watch Related Video in #geneticteacher) #geneticteacher

(PCR-RAPD Marker) Initial denaturation start at 94 degree Celsius for 10 minutes and denaturation temperature should be at 94 degree Celsius for 1 minute and annealing temperature should be at 37 to 45 degree Celsius for1 minute and extension temperature should be at 72 degree Celsius for 1 minute and final extension should be at 72 degree Celsius for 10 minutes and cooling temperature should be at 4 degree Celsius forever and Polymorphic band must be clone and make sequence to design new primers to amplify specifically (Watch Related Video in #geneticteacher) #geneticteacher

Restriction Maps are position of restriction sites within genome (For example DNA extraction from disease and resistant plants then digested by restriction enzymes and resolved by gel electrophoresis and the gel that digested by Hind III has monomorphic pattern whereas the gel that digested by Eco-RI has polymorphic pattern), Restriction enzymes that have shorter sequence have greater number of fragments (Watch Related Video in #geneticteacher) #geneticteacher

Restriction Fragment Length Polymorphism or RFLP marker require pure and high molecular weight of DNA to digest DNA with restriction enzymes and detect digested DNA by southern blot technique and detect interested gene by PCR, By making DNA extraction and digest DNA with restriction enzymes or restriction endonucleases then make gel electrophoresis of DNA fragments after that transfer DNA fragments using southern blotting then hybridizing DNA with label probe and make autoradiography, RFLP marker is co-dominant inheritance because it estimate Heterozygosity and multi loci specific marker because it define two or more alleles and it isn't require prior sequence information, On the other hand RFLP marker require large amount of DNA and present low level of polymorphism in some species and present few loci per assay with time consuming and cost, RFLP marker use in genetic diversity and genetic relationship and genetic drift and selection and whole genomic mapping and gene tagging, DN...

Restriction Enzymes or Restriction Endonucleases or Molecular Scissors found in bacteria to provide defense mechanism against invading viruses by degrading viral DNA without affecting bacterial DNA, Each bacterium has its own unique restriction enzyme and each enzyme recognize only one type of sequence so DNA sequence that recognize by restriction enzymes called palindromes which are sequences read same on two strands in opposite directions, Mechanisms of restriction enzymes start by scan DNA then bind to DNA to recognize specific sequence after that cut each sugar-phosphate backbone of DNA by hydrolyze phosphodiester bonds, Restriction Enzymes have four types: (Type-1: Cleave DNA at random length from recognition sites) (Type-2: Cleave DNA in close of recognition site so it useful for gene analysis and cloning) (Type-3: Cleave outside recognition sequence so it require two recognition sequence in opposite directions within same DNA) (Type-4: Cleave only modified DNA in approximate 30 ...

Restriction enzymes used in gene characterization by RFLP markers and gene mapping such as restriction maps and gene manipulation such as gene cloning and protein expression experiments (Watch Related Video in #geneticteacher) #geneticteacher

For Restriction Enzymes Mapping must digest DNA with restriction enzymes then resolve digested fragments by gel electrophoresis and the resulted bands on gel refer to., number of restriction sites while size of bands refer to distance between restriction sites (Watch Related Video in #geneticteacher) #geneticteacher

Sticky 5’ Overhang, Sticky 3’ Overhang, Blunt Ends without Overhang, Sticky ends Restriction Enzymes called also Cohesive ends Restriction Enzymes because they are useful for DNA manipulation and can be converted to blunt ends with nuclease or polymerase while Blunt ends Restriction Enzymes useful for certain types of DNA cloning experiments and can be converted to sticky ends by ligating to synthetic adaptors (Watch Related Video in #geneticteacher) #geneticteacher

Restriction Enzymes have four types: (Type-1: Cleave DNA at random length from recognition sites) (Type-2: Cleave DNA in close of recognition site so it useful for gene analysis and cloning) (Type-3: Cleave outside recognition sequence so it require two recognition sequence in opposite directions within same DNA) (Type-4: Cleave only modified DNA in approximate 30 base pair away from sites) (Watch Related Video in #geneticteacher) #geneticteacher

Mechanisms of restriction enzymes start by scan DNA then bind to DNA to recognize specific sequence after that cut each sugar-phosphate backbone of DNA by hydrolyze phosphodiester bonds (Watch Related Video in #geneticteacher) #geneticteacher

Restriction Enzymes or Restriction Endonucleases or Molecular Scissors found in bacteria to provide defense mechanism against invading viruses by degrading viral DNA without affecting bacterial DNA, Each bacterium has its own unique restriction enzyme and each enzyme recognize only one type of sequence so DNA sequence that recognize by restriction enzymes called palindromes which are sequences read same on two strands in opposite directions (Watch Related Video in #geneticteacher) #geneticteacher

Probe Sequence usually range from 100 to 500 base pair in length to detect target region in the single strand of DNA, Sequence Polymorphism use to make alignment between sequence queries to detect single nucleotide polymorphism, Length Polymorphism refer to number of repeated sequence across genome (Watch Related Video in #geneticteacher) #geneticteacher