Paddg rice anti reverse related gene trehalose-6-phosphoric acid phosphorylase gene and its application
A technology of phosphorylase and trehalose, applied in the field of botany, can solve the problems of short plants and slow plant growth
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0081] Cloning of Rice Trehalose-6-Phosphate Phosphorylase Gene
[0082] Using a 10,000-point cDNA microarray representing approximately 9,000 rice genes (purchased from Shanghai United Gene Boxing Gene Chip Co., Ltd.), the salt-tolerant rice variety "Nona Bokra" (available from the rice variety resource of the International Rice Research Institute, etc. library) under 150mM NaCl high-salt stress, a series of high-salt-responsive genes were identified. Among them, a fast-response trehalose-6-phosphate phosphorylase gene OsTPPA was identified 20 minutes after salt stress.
[0083] The Nona OsTPPA gene of the full-length ORF was cloned by screening the genome library and cDNA library of the salt-resistant variety Nona Bokra under salt stress. The method is as follows: the cDNA library of the highly salt-tolerant rice variety NonaBokra was constructed by conventional methods. Using the cDNA library as a template, the PCR method was used to obtain the amplified product, which wa...
Embodiment 2
[0086] Rice trehalose-6-phosphate phosphorylase is a salt tolerance-related protein
[0087] Use the oligonucleotides corresponding to the first 20bp and the last 20bp of ORF in SEQ ID NO: 1 as primers (or amplified products as probes), and OsTPPA gene in resistant and susceptible varieties by conventional Northern and RT-PCR techniques The expression patterns under moderate salt stress were analyzed.
[0088] The results showed that the expression of OsTPPA in the roots and shoots of the salt-tolerant rice variety Nona Bokra could be rapidly induced by salt, drought and cold treatments. After Nona Bokra was salt-treated for 20 minutes, OsTPPA was up-regulated in seedlings, but down-regulated in the salt-sensitive variety IR28 (available from the rice variety resource bank of the International Rice Research Institute, etc.). This indicates that OsTPPA is related to salt tolerance and plays an important role in rice stress resistance.
Embodiment 3
[0090] Preparation of rice OsTPPA transgenic material
[0091] This embodiment adopts the binary vector pHB (pHB is that the GUS gene in the pCAMBIA3301 plasmid purchased from CAMBIA Company is replaced by the hygromycin resistance gene (Hygr ) obtained plasmid) as rice transgene vector. The vector encodes a bacterial origin of replication (ori), a kanamycin resistance gene (Kanr), a hygromycin resistance gene (Hygr), a herbicide resistance gene (Bar), a CaMV35S promoter, and a termination signal for the NOS gene sequence and the restriction enzyme cloning site (MCS) between the latter two. The cDNA of OsTPPA can be inserted forward or reverse into the restriction endonuclease cloning site to construct a transgenic plasmid.
[0092] (1) Construction of transgenic plasmids for positive sense overexpression of OsTPPA:
[0093]In this embodiment, the constructed salt stress cDNA library derived from the salt-resistant variety Nona Bokra was used as a template, and the PCR olig...
PUM
Login to View More Abstract
Description
Claims
Application Information
Login to View More