Which of the following is a unique attribute of CRISPR/Cas9 compared to restriction enzymes Group of answer choices it can be targeted to any specific region of the genome as long as there is an adjacent PAM motif it was first discovered in bacterial cells it can cut double-stranded DNA sequences it can be used for molecular cloning experiments
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- Cloning Genes Is a Multistep Process The following DNA sequence contains a six-base sequence that is a recognition and cutting site for a restriction enzyme. What is this sequence? Which enzyme will cut this sequence? (See Figure 13.5 for help.) 5 CCGAGGAAGCTTAC 3 3 GGCTCCTTCGAATG 5What is the role of the following molecules in the CRISPR mechanism? * Sequence of 3 nucleotide Not a RNA significant sequence Endonuclease motif that is Host genetic synthesized role in information to match a a binding CRISPR target site mechanism sequence Cas9 guide RNA DNA polymerase PAM Cellular DNA TRNA O O 口 ロ 口 ロロWhich of the following would you need to clone a human gene? Choose all that apply A restriction enzyme Human DNA A donor goat nucleus An enucleated egg A surrogate mother DNA ligase A plasmid vector (or other type of DNA vector, e.g. phage, BAC)
- Which of the following are required to set up a CRISPR-Cas9 experiment to fix a mutation in a gene so that the gene sequence will be changed to the wild-type sequence? Circle all that apply. O CRISPR-Cas9 nuclease O sgRNA (single guide RNA) complementary to the part of the target sequence to be fixed O a PAM site (immediately adjacent to the target sequence in the genome) O donor DNA containing wild-type sequence of the part of the gene that is targeted (donor DNA can also be called repair template)Please place the stages of a CRISPR-cas9 gene editing workflow in the correct order below. Note, there is one incorrect and state why? Sequence Select for correctly gene edited cells (using antibiotic resistance and/or colour production for example) Clonal isolation Synthesise DNA insert oligonucleotides Clonal characterisation (analysis of phenotype) Add green fluorescent protein gene sequence into plasmid to aid selection of correctly transfected cells. Transfect cells Clone into CRISPR-cas9 expression vector Purify plasmids Design a set of targeting sequences using algorithms.CRISPR/Cas9 can be used in genome editing. Among the following statements, which one is correct? The CRISPR and the Cas9 parts both recognise the DNA target sequence and then recruit an endonuclease for cutting it The Cas9 part recognises the DNA target sequence and the CRISPR part cuts it The CRISPR and the Cas9 parts both recognise and cut the DNA target sequence. The CRISPR part recognises the DNA target sequence and Cas9 cuts it.
- Place the stages of a CRISPR-cas9 gene editing workflow in the correct order (there is one incorrect stage) Design a set of targeting sequences using algorithms. Clonal isolation Transfect cells Synthesise DNA insert oligonucleotides Purify plasmids Sequence Clonal characterisation (analysis of phenotype) Select for correctly gene edited cells (using antibiotic resistance and/or colour production for example) Add green fluorescent protein gene sequence into plasmid to aid selection of correctly transfected cells. Clone into CRISPR-cas9 expression vectorLabel the following parts of the CRISPR-Cas 9 system. C A B C D Target DNA strand PAM sequence Cas9 protein Single-guide RNADuring nucleic acid hybridization, the probe is labelled for DNA stability to increase probe-test DNA binding to identify the location of probe and the test DNA binding for amplification
- Which of the following is required to make complementary DNA (CDNA) from RNA? reverse transcriptase DNA ligase DNA ligase gene cloning restriction enzymes (endonucleases)You want to edit the cystic fibrosis gene using the CRISPR-cas9 system. In order to target your gene of interest to make the cut, which of the following components of the CRISPR-cas9 system should be specific to the cystic fibrosis gene? All of these components of the CRISPR system need to be specific to the gene of interest The editing template The guide RNA The CRISPR associated proteinCRISPR-cas9 is a powerful technique for gene editing. Please place the stages of a CRISPR-cas9 gene editing workflow in the correct order below options are on the right