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Identifying Sweet Orange (Citrus Sinensis (L.) Osbeck) Germplasm Accessions with Inter-simple Sequence Repeat (ISSR) Markers

Author: HuangXiaoDan
Tutor: YingTieJin
School: Zhejiang University
Course: Of Food Science
Keywords: Sweet orange ISSR Molecular markers Germplasm
CLC: S666.4
Type: Master's thesis
Year: 2006
Downloads: 308
Quote: 11
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Abstract


22 In this study, the Chongqing Institute of Pomology sweet orange (Citrus sinensis (L.) Osbeck) sample material , the use of ISSR markers to germplasm identification . The main findings are as follows: ( 1) to obtain a time-saving , simple and suitable for ISSR-PCR amplification of the sweet orange genome DNA extraction method . Relatively short time required to extract the sweet orange genome DNA through the law , and do not need repeated phenol : chloroform: isoamyl alcohol extraction , extracted DNA OD 260/280 1.65 - 1.90 range ; (2) applies to the laboratory by the annealing temperature , MgCl 2 < / sub> and dNTP concentrations test , get ISSR-PCR reaction system and amplification procedures are as follows : PCR reaction system : 20μL of the amplification reaction mixture comprises 10 × PCR buffer , 2.2mmol / L mM MgCl 2 , dNTP each concentration for 220μmol / L , 1μmol / L primer 2 % formamide , 1 unit Taq enzyme ( Shanghai Shenggong ) , and 25ng DNA template ; PCR program : 94 ° C denaturation 6min ; 27-35 cycles , 30s ,54-58 ℃ annealing 45s at 94 ℃ , 72 ℃ extension of 2min (Touch-down PCR); 72 ℃ for 7min. Wherein the annealing temperature for primer rather isobutyl ; (3) from 22 ISSR primer was Filter 13 can obtain efficient amplification of the primer was and Trials of annealing temperatures and cycle times , create applies to these 13 ISSR primer was the PCR program . These 13 primers were used for the 22 oranges sample ISSR-PCR amplification, to obtain a total of 218 strips , the strip size within the range of from 100-3000bp , wherein 11 samples amplified by eight primer 15 polymorphic bands , these 11 samples from the 22 sample areas separately . The results showed that the closer relationship between all varieties of the sweet orange ; (4) can be found a possible link between quality analysis and molecular marker studies compared , but not by the experimental the exact relationship between results obtained .

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CLC: > Agricultural Sciences > Gardening > Fruit trees gardening > Citrus > Orange
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