The method of claim 26, wherein the at least one gRNA sequence comprises a first gRNA sequence that is complementary to a first DNA segment of the target nucleic acid and a second gRNA sequence that is complementary to a second DNA segment of the target nucleic acid.ģ3. The method of claim 26, wherein the target nucleic acid is a DNA sequence in a cell.ģ2. The method of claim 26, wherein elements (i), (ii), and (iii) are packaged into a single vector.ģ1. A method of inserting a transposon into a target site of a target nucleic acid to disrupt expression of the target nucleic acid, the method comprising providing to the target nucleic acid (i) a fusion protein of claim 1, and (ii) at least one gRNA sequence complementary to a segment of a target nucleic acid, wherein the segment is adjacent to the target site to direct transposon insertion, and, optionally, (iii) at least one mini-transposon.Ģ7. The system of claim 17, wherein the at least one gRNA sequence comprises a first gRNA sequence that is complementary to a first DNA segment of the target nucleic acid and a second gRNA sequence that is complementary to a second DNA segment of the target nucleic acid.Ģ6. The system of claim 22, wherein the transposon end sequence comprises a sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQ ID NO:9, or reverse complement thereof, or SEQ ID NO:12, or a reverse complement thereof.Ģ5. The system of claim 21, wherein the transposon end sequence comprises an inverted repeat of a Himar1 transposon or Tn5 transposon.Ģ4. The system of claim 21, wherein the mini-transposon comprises a payload sequence comprising a 5′ and 3′ end, a first transposon end sequence that is fused to the 5′ end of a payload sequence and a second transposon end sequence that is fused at the 3′ end of the payload sequence.Ģ3.
The system of claim 17, further comprising at least one mini-transposon.Ģ2. A system comprising a fusion protein according to claim 1 and at least one gRNA sequence complementary to a segment of DNA sequence, wherein the segment is adjacent to a target site of a target nucleic acid.Ģ1. The fusion protein of claim 14, wherein the fusion protein comprises one or more mutations selected from the group consisting of M470_I476del, A471_I476del, and S458A.ġ7. The fusion protein of claim 1, wherein the fusion protein comprises a polypeptide sequence comprising at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO:5.ġ5. The fusion protein of claim 10, wherein the fusion protein comprises one or more mutations selected from the group consisting of Y12A, Y12S, F31A, W119A, V120A, P121A, R122A, E123A, and L124A.ġ4. The fusion protein of claim 1, wherein the fusion protein comprises a polypeptide sequence comprising at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO:3.ġ1. The fusion protein of claim 5, wherein the Cas9 protein is catalytically dead.ġ0. The fusion protein of claim 1, wherein the Cas protein is Cas9.Ħ. The fusion protein of claim 1, wherein the transposase comprises a polypeptide sequence comprising at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to the amino acid sequence of SEQ ID NO: 1, or 4, or active fragments thereof.ĥ.
A fusion protein comprising a transposase fused to a Cas protein, wherein the transposase is Himar1 or Tn5.ģ.