	US 12,227,752 B1
	*Use of MTNAC33 gene and an encoded protein thereby in improving the biomass and drought tolerance of Medicago sativa* L.**
Chunxiang Fu, Qingdao (CN); Ruijuan Yang, Qingdao (CN); Wenwen Liu, Qingdao (CN); Zhaoming Wang, Qingdao (CN); Feng Yuan, Qingdao (CN); Ying Sun, Qingdao (CN); Shiqie Bai, Qingdao (CN); Chuanen Zhou, Qingdao (CN); Jiajun Yan, Qingdao (CN); Yaling Liu, Qingdao (CN); Xiaowei Zhang, Qingdao (CN); and Shanshan Jiang, Qingdao (CN)
Assigned to QINGDAO INSTITUTE OF BIOENERGY AND BIOPROCESS TECHNOLOGY, CHINESE ACADEMY OF SCIENCES, Qingdao (CN); and Inner Mongolia Pratacultural Technology Innovation Center Co. Ltd, Hohhot (CN)
Filed by QINGDAO INSTITUTE OF BIOENERGY AND BIOPROCESS TECHNOLOGY, CHINESE ACADEMY OF SCIENCES, Qingdao (CN); and Inner Mongolia Pratacultural Technology Innovation Center Co. Ltd, Hohhot (CN)
Filed on Jul. 25, 2024, as Appl. No. 18/783,627.
Int. Cl. C12N 15/82 (2006.01); A01H 1/04 (2006.01)

CPC C12N 15/8273 (2013.01) [A01H 1/04 (2013.01); C12N 15/8205 (2013.01)]

3 Claims

1. A method for improving biomass yield and drought tolerance of alfalfa, comprising the following steps:

(1) constructing a pEG100-MtNAC33 vector;

(2) introducing a MtNAC33 expression cassette comprising the pEG100-MtNAC33 vector into alfalfa by using an ultrasonic-assisted leaf disc transformation procedure mediated by an Agrobacterium tumefaciens strain EHA105; wherein the pEG100-MtNAC33 vector contains an MtNAC33 gene having the nucleotide sequence shown in SEQ ID NO: 1; and

(3) conducting DL-Phosphinothricin resistance selection to obtain a positive transgenic alfalfa plant; wherein the positive transgenic plant shows delayed flowering time, increased leaf-to-stem ratio, improved dry biomass and increased drought tolerance.