Tomato SlML1 gene and application

A gene, tomato technology, applied in the field of tomato SlML1 gene

Inactive Publication Date: 2015-04-29
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Before the present invention was published, there was no publication or report on the functional re

Method used

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  • Tomato SlML1 gene and application
  • Tomato SlML1 gene and application
  • Tomato SlML1 gene and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Construction of tomato cDNA library

[0042] 1. Isolation and Detection of Tomato Total RNA

[0043] Take 2 g of fresh tomato (tomato S. pennellii) leaves, quickly grind them into powder with liquid nitrogen in a mortar, and quickly transfer to 10 mL of extraction buffer (CTAB (W / V) 2%, Tris-HCl) preheated at 65 °C (pH 8.0) 100mmol L -1 , EDTA25mmol·L -1 , NaCl2.0mol L -1 , PVP402%, spermidine 0.5g / L, mercaptoethanol 2%), shake and mix well; extract twice with equal volume of chloroform, and centrifuge at 7500g for 15 minutes. Add 1 / 4 volume of 10M LiCl to the supernatant, mix well and place it at 4°C to precipitate overnight; centrifuge at 7500g for 20 minutes, and use 500 μL SSTE (SDS0.5%, NaCl1mol L -1 , Tris-HCl (pH8.0) 10mmol L -1 , EDTA1mmol·L -1 , dissolved at 65°C for 5 minutes. Extract with an equal volume of chloroform, centrifuge at 13,000g for 5 minutes; add 2 times the volume of absolute ethanol to the supernatant, and store at -70°C for 2 hours; cen...

Embodiment 2

[0047] Cloning of related genes in tomato

[0048] Primers P1: 5'-ATGGGAAGATCACCAT-3', P2: 5'-T TAAAATACTGATGAACCATCAAC-3' were designed according to the tomato genome sequence, and the PCR reaction was carried out using tomato cDNA as a template. The reaction system was as follows: Taq buffer 2.5 μL, MgCl 2 (25mM) 1.8μL, dNTP (2.5mM) 1μL, P1 primer (10pmol) 1μL, P2 primer (10pmol) 1μL, Taq enzyme 0.4μL. The PCR reaction conditions were 5 minutes of pre-denaturation at 94°C, 40 seconds at 94°C, 40 seconds at 50°C, 4 minutes at 72°C, 35 cycles of extension at 72°C for 10 minutes, and storage at 4°C. Take 7ul of PCR product and add 3ul of bromofinland for agarose gel electrophoresis, take a picture half an hour later, observe the gel map, select the amplified product with a single band and the correct position and send it to Huada Gene Company for sequencing, and the sequence alignment is correct. Tomato tomato SlML1 gene.

Embodiment 3

[0050] Bioinformatics analysis of SlML1 gene

[0051] The length of the obtained full-length cDNA of tomato S1ML1 is 1005bp, the sequence is shown in SEQ ID NO.1, wherein the open reading frame is located at 1-1005bp, and the sequence of the encoded protein is shown in SEQ ID NO.2. The full-length tomato cDNA sequence was searched for nucleotide homology in Non-redundant GenBank+EMBL+ DDBJ+PDB and Non-redundant GenBank CDS translation+PDB+Swissprot+Super date+PIR databases using BLAST program. Compared with R1R3-MYB of other species, the gene is highly conserved at the amino acid level and has typical MYB_DNA-bind and SANT domains. Such as figure 1 .

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Abstract

The invention discloses a tomato SlML1 gene and application, and belongs to the field of biological techniques. The cDNA sequence and the encoding amino acid sequence of the gene are as shown in SEQIDNO.1 and SEQIDNO.2. The transgenosis experiment shows that the gene not only has the functions of improving the quality and increasing the yield of tomatoes, but also has the properties of improving the drought resistance, the fungal disease resistance, bacterial disease resistance and TMV and insect resistance of tomatoes, and moreover the shelf life of tomatoes can be prolonged. Therefore, SlML1 can be used as a target gene introduction plant for improving the quality and the resistance and increasing the yield of plants and has good application prospect in breeding, quality improvement and resistance study on tomatoes.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a tomato S1ML1 gene, and also relates to the application of the gene in tomato strain quality, biotic stress and abiotic stress resistance and shelf life. Background technique [0002] The MYB transcription factor family refers to a class of transcription factors containing the MYB domain. The MYB domain is a DNA-binding domain containing 52 amino acids. MYB transcription factors in plants have a common feature of a MYB domain consisting of about 52 amino acids at their N-terminus. Clorless1 (C1) of maize is the first MYB transcription factor isolated and identified in plants (Paz-Areset et al., The regulatory c1 locus of Zea may encodes a protein with homology to myb proto-oncogene products and with structural similarities to transcriptional activators.EMBO J,1987,6:3553-3558), subsequently, Arabidopsis (Romero et al.,More than80R2R3-MYB regulatory genes in the genome ...

Claims

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Application Information

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IPC IPC(8): C12N15/29C07K14/415C12N15/84A01H5/00
Inventor 罗杰默罕默德·艾瓦斯刘贤青张红艳
Owner HUAZHONG AGRI UNIV
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