Modified excisable DAS59122-7 maize transgenic locus
Transgenic INIR4 maize plants comprising modifications of the DAS59122-7 maize locus which provide for facile excision of the modified DAS59122-7 transgenic locus or portions thereof, methods of making such plants, and use of such plants to facilitate breeding are disclosed.
1. A transgenic maize plant cell comprising an INIR4 transgenic locus comprising an originator guide RNA recognition site (OgRRS) in a first DNA junction polynucleotide of a DAS59122-7 transgenic locus and a cognate guide RNA recognition site (CgRRS) in a second DNA junction polynucleotide of the DAS59122-7 transgenic locus, wherein said INIR4 transgenic locus comprises the DNA molecule set forth in SEQ ID NO: 27 or an allelic variant thereof.
2. The transgenic maize plant cell of claim 1 , wherein said INIR4 transgenic locus comprises the DNA molecule set forth in SEQ ID NO: 27.
3. A transgenic maize plant part comprising the maize plant cell of claim 1 .
4. The transgenic maize plant part of claim 3 , wherein said maize plant part is a seed.
5. A transgenic maize plant comprising the maize plant cell of claim 1 .
6. A method for obtaining a bulked population of inbred seed comprising selfing the transgenic maize plant of claim 5 and harvesting seed comprising the INIR4 transgenic locus from the selfed maize plant.
7. A method of obtaining hybrid maize seed comprising crossing the transgenic maize plant of claim 5 to a second maize plant which is genetically distinct from the first maize plant and harvesting seed comprising the INIR4 transgenic locus from the cross.
8. A DNA molecule comprising SEQ ID NO: 27.
9. A processed transgenic maize plant product comprising the DNA molecule of claim 8 .
10. A biological sample containing the DNA molecule of claim 8 .
11. A method of detecting a maize plant cell comprising the INIR4 transgenic locus of claim 1 , comprising the step of detecting a DNA molecule comprising SEQ ID NO: 10.
12. A method of excising the INIR4 transgenic locus from the genome of the maize plant cell of claim 1 , comprising the steps of:
(a) contacting the INIR4 transgenic locus of the plant cell with: (i) a Cas12 RNA dependent DNA endonuclease (RdDe); and (ii) a guide RNA (gRNA) capable of hybridizing to the guide RNA hybridization site of the OgRRS and the CgRRS of SEQ ID NO: 27; wherein the Cas12 RdDe recognizes a OgRRS/gRNA and a CgRRS/gRNA hybridization complex; and,
(b) selecting a transgenic plant cell, transgenic plant part, or transgenic plant wherein the INIR4 transgenic locus flanked by the OgRRS and the CgRRS has been excised.
13. The method of claim 12 , wherein the guide RNA comprises an RNA sequence encoded by SEQ ID NO: 13.