Molecular marker method for identifying tomato yellow leaf curl virus resistance gene Ty-1
A technology of tomato yellowing curved leaves and molecular markers, applied in the field of agricultural biology, can solve the problems of low linkage degree of marker genes, many operation procedures, and the need for enzyme digestion, etc., to achieve simple electrophoretic band patterns, improve accuracy, and speed up The effect of the breeding process
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Embodiment 1
[0021] Example 1, Screening of Molecular Marker Primers
[0022] The BSA method was used to construct the disease-resistant gene pool and the disease-susceptible gene pool for the screening of molecular marker primers.
[0023] Zamir et al. located the yellow leaf curl virus resistance gene Ty-1 on chromosome 6, and the RAPD marker and CAPS marker linked to this gene have been screened out. According to the positioning result, tomato genome sequence and SSR genetic map, the present invention newly designs a series of marker primers. Then, polymorphism screening was carried out for the designed series of marker primers against disease-resistant materials and non-disease-resistant materials. As a result of the screening, only one pair of primers can amplify the polymorphism, which can be used as molecular marker primers. The molecular marker primer can amplify a 690bp fragment in the disease-resistant material containing the Ty-1 site; it can amplify a 409bp fragment in the no...
Embodiment 2
[0026] Example 2, identification of yellow leaf curl virus resistance gene using screened molecular marker primers
[0027] 1. Extraction of tomato genomic DNA by CTAB method
[0028] The materials to be identified are tomato materials of the following varieties: Shenfen 8, Shenfen 918, Puhong 9, Puhong 10, Shenfen V-1, Dongnong 721, Xianzheng Hongzhu.
[0029] Collect the new leaves of the tomato material of the above-mentioned varieties, and extract DNA with the CTAB method. The specific extraction method is as follows:
[0030] (1) Take 1-50g of fresh plant material and grind it into powder in liquid nitrogen. (2) Transfer the frozen powder into a pre-cooled centrifuge tube, immediately add an equal volume (w / v) of 2×CTAB extraction buffer, and incubate at 65°C for 10-20 minutes, shaking from time to time. (3) Add an equal volume of chloroform / isoamyl alcohol, gently invert the centrifuge tube to mix, and centrifuge at 12000 r / min for 10-20 minutes at room temperature. (...
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