Question

To differentiate between the host genome and the pathogen to target for elimination in the CRISPR-Cas 9 system

	a.	a protospacer adjacent motif (PAM) must follow the protospacer
	b.	the spacers must be interspersed with CRISPR
	c.	the spacers must be sequenced in order of previous contact with a pathogen
	d.	the spacer regions must base pair with target sequences

CRISPR-Cas gene editing systems require a series of sequences that permit mutation, knock-out, or knock-in capabilities. Select the required sequence configuration for optimal editing results.

	a.	pre-crRNA; tracrRNA; Cas1Cas 2 identifies protospacer; CRISPR-protospacer inserted in front of CRISPR-spacer sequence; Cas-9
	b.	CRISPR-protospacer inserted in front of CRISPR-spacer sequence; Cas1Cas2 identifies protospacer; pre-crRNA; tracrRNA; Cas-9
	c.	Cas1Cas2 identifies protospacer; CRISPR-protospacer inserted in front of CRISPR-spacer sequence; pre-crRNA; tracrRNA; Cas-9
	d.	pre-crRNA; tracrRNA; Cas1Cas 2 identifies protospacer; Cas-9; CRISPR-protospacer inserted in front of CRISPR-spacer sequence

Answer 1

Answer:

Q1) OPTION A) A protospacer adjacent motif (PAM) must follow protospacer

Rationale:

• CRISPR cas system is an adaptive immune system of many bacteria from invading viruses.
• PAM is a short sequence motif which is located adjacent to the protospacer (invading DNA).
• This PAM is important for diifferentiating and recognizing the invading DNA (non self) from self DNA.
• After differentiating it is involved in cleavage and elimination of invading DNA.

Q2) Option C)Cas1Cas2 identifies protospacer; CRISPR-protospacer inserted in front of CRISPR-spacer sequence; pre-crRNA; tracrRNA; Cas-9

CRISPR Cas system follows three phase,

1. First phase is adaptation stage,in which an enzyme complex Cas 1- Cas2 recognises the invading viral DNA.After recognising it acquires a region called protospacer.This protospacer is then inserted into the CRISPR spacer array and in between the repeats.
2. After adaptation expression and maturation stage ,in this stage once the protospacer is integrated in to the CRISPR ,the locus is transcribed into a precusor RNA(pre CRISPR RNA).After this pre CRISPR RNA again transcribed in to mature trans acting CRISPR RNA( tracrRNA).
3. Third phase is Interference phase,here cas proteins reaches the tracrRNA and form an effector complex called crRNA-Cas RNP complex(in Type II CRISPR cas system tracrRNA-Cas9 complex forms).tcrRNA maintains cas 9 in active form to traget the foreign DNA with the help of PAM.These effectot complex have complementary sequence against the protospacer,which then binds with it and cause endonucleolytic cleavage of protospacer.

Hope this helps you:)