Modified excisable 5307 maize transgenic locus lacking a selectable marker
Transgenic INIR17 maize plants comprising an ecry3.1Ab expression cassette linked to a secondary nopaline synthase terminator element which lack a selectable marker gene and/or which comprise modifications that provide for facile excision of the INIR17 transgenic locus from the maize plant genome are provided. Genomic DNA of INIR17 transgenic plants, detection of INIR17 plants and products thereof, methods of making INIR17 plants, and use of INIR17 plants to facilitate breeding are disclosed.
1. A transgenic maize plant cell comprising an INIR17 transgenic locus comprising a cognate guide RNA recognition site (CgRRS) in a 5′ junction polynucleotide of a 5307 maize event transgenic locus, an originator guide RNA recognition site (OgRRS) in a 3′ junction polynucleotide of the 5307 maize event transgenic locus, and a deletion of the 1 st NOS terminator, the ZmUbiInt promoter, and the phosphomannose isomerase coding region of the 5307 maize event transgenic locus, wherein said INIR17 transgenic locus comprises the DNA molecule set forth as SEQ ID NO: 33 or an allelic variant thereof.
2. The transgenic maize cell of claim 1 , wherein the INIR17 transgenic locus comprises the DNA molecule of SEQ ID NO: 33.
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 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 INIR17 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 INIR17 transgenic locus from the cross.
8. A DNA molecule comprising SEQ ID NO: 33.
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 transgenic locus of claim 1 , comprising the step of detecting a DNA molecule comprising SEQ ID NO: 16 or 40.
12. A method of excising the INIR17 transgenic locus comprising a DNA molecule of SEQ ID NO: 33 or an allelic variant thereof from the genome of the maize plant cell of claim 1 , comprising the steps of:
(a) contacting the genome of the maize 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 originator guide RNA recognition site (OgRRS) and the cognate guide RNA recognition site (CgRRS) of the transgenic locus; 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 transgenic locus flanked by the OgRRS and the CgRRS has been excised.