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1.
A random amplified polymorphic DNA (RAPD) marker named OPC06-1900 was previously found linked to a fertility restorer gene (Rfw) for cytoplasmic male sterility (CMS) in radish (Raphanus sativus L.). The RAPD marker was converted to a dominant sequence characterized amplified region (SCAR) marker SCC06-1894 by molecular cloning and nucleotide sequencing. A BLAST search revealed that the SCAR marker SCC06-1894 showed significant homology to the corresponding regions of Arabidopsis and Brassica sulfate transporter genes. The presence of the intron and exon of the DNA fragment SCC06-1894 was demonstrated by comparing RT-PCR and PCR products. Thus, allele-specific oligonucleotide primers were designed to amplify the SCAR marker SCC06-415. PCR test with F2 plants and sequence analysis showed that SCC06-1894 and SCC06-415 were allelic, linked to Rfw/rfw gene at 8.0 cM. Nine oligonucleotide primers were designed based on a single radish nuclear restorer gene mRNA. A survey of these primer combinations by bulked segregant analysis (BSA) identified three polymorphisms. The three PCR-based markers were co-segregant in the coupling phase and distant from the Rfw gene by 1.4 cM. These specific markers distributed on both sides of the Rfw gene and will be helpful for breeding new rapseed (Brassica napus L.) restorer lines.  相似文献   

2.
The Rfo fertility restorer gene for the Ogura cytoplasmic male sterility (CMS) applied for oilseed rape hybrid seed production can be monitored with the use of the RAPD OPC021150 marker while molecular breeding. The aim of this work was to convert the RAPD marker into a more suitable SCAR marker. Total DNA was isolated from a doubled haploid line derived from the line BO20 (INRA, France). A fragment of 1150‐bp linked to the Rfo gene was PCR amplified with the use of the RAPD OPC02 primer, cloned and sequenced. A pair of primers was designed and PCR amplification was performed to develop a SCAR marker for the Rfo gene. The new marker was applied for analysis of 220 oilseed rape lines comprising doubled haploid and inbred restorer lines, restored hybrids as well as F1 and F2 recombinant generations involving restorer lines. Simultaneously, the RAPD OPC02 marker was used and it revealed that the markers are equivalent to each other. However, the developed new SCAR marker has made the analysis more practical, rapid and efficient.  相似文献   

3.
利用42条RAPD（Random Amplified Polymorphic DNA）随机扩增引物分析工业大麻品种“火麻一号”组成的雄性或雌性DNA池（DNA pools）,结果显示,引物OPV-08扩增得到一条大小为869bp与工业大麻雄性相关的特异条带。根据测序结果,合成了两条SCAR（Sequence Characterized Amplified Region）标记引物,该SCAR标记不仅可以对工业大麻雌雄异株材料花期已知性别的雌雄植株进行准确鉴定,还可以对幼苗期未知性别的大麻雌雄植株进行鉴定;也可对雌雄同株材料可能出现的雄化进行早期鉴定。这不仅为工业大麻早期性别鉴定提供基础,且为减少雌雄同株材料的雄化提供支撑。  相似文献   

4.
There is an urgent need for early sex identification to support field planting in Ginkgo biloba L., due to the different economic and medicinal values between male and female trees. An easy, rapid and reliable molecular method for sex type determination of G. biloba was reported in the paper. Random amplification of polymorphic DNA (RAPD) and sequence-characterized amplified region (SCAR) were used to search for specific molecular markers linked to the sex locus. A total of 48 primers were used for screening of specific RAPD markers in six male and three female samples. Only one primer, S10, showed different amplification band patterns associated with sex types. Then the sex-specific bands, S10-BandA and S10-BandB, were cloned and sequenced. Based on the sequences two pairs of SCAR primers, GBA and GBB, were designed. The GBA primers amplify a single 571 bp band in male samples but not in female samples, and DNA amplification using GBB primers could generate a 688 bp band only in the female individuals. Finally, the SCAR primers were used to test 16 sex-unknown samples. SCAR primers developed in this paper can be used as effective, convenient and reliable molecular markers for sex identification in G. biloba.  相似文献   

5.
棉花抗卷叶病RAPD和SCAR标记研究   总被引：3，自引：0，他引：3  
利用RAPD对抗卷叶病品系CNH 123、CNH 1012和感病品系CNH 1020、CNH 120分别建立了抗、感病多态带,用80个引物进行PCR扩增.引物OPC 02在抗病品系CNH 123和CNH 1012中得到1700 bp的特征片段,建立了10个抗病及感病DNA库并进行混合分组分析,在F2群体用OPC 02重现了上述特征片段,并将该片段改造为SCAR标记,设计引物对为5'-GT-GAGGCGTCAGAGGGAT-3'(正向)and 5'-GTTGCCGTGCACTAGGCT-3'(反向).利用Mapmaker软件得到10个F2分离群体的RAPD分离图谱.  相似文献   

6.
Sex identification in Pistacia species during the long juvenile stage is an economically desirable objective. Due to the lack of morphological methods to identify sex at this stage, the application of molecular markers is expected to facilitate breeding programs. The aim of our study was to identify a marker closely linked to sex loci in Pistacia atlantica Desf subsp. mutica, P. khinjuk, and P. vera subsp. Sarakhs. Samples were collected from both male and female plants of each species, and their band patterns were analyzed according to the presence or absence of specific bands. Thirty random amplified polymorphic DNA (RAPD) primers and a pair of sequence characterized amplified region (SCAR) primers were tested as potential markers of sex in wild Pistacia species. Among the RAPD primers, only BC1200 was found to amplify a specific sex band present in female plants. Based on our analysis of all individual samples, a fragment of approximately 300 bp was amplified in female trees but absent in male ones. Although sex determination mechanisms in Pistacia are still unknown, they may be controlled by a single locus that acts as a trigger. The SCAR technique has proved to be a reliable technique in gender determination of pistachio genotypes at the seedling phenophase. This method could reduce both the time and costs associated with breeding programs.  相似文献   

7.
DNA from female and male hemp (Cannabis sativa L.) plants belonging to nine different varieties were screened with180 RAPD primers in a search for sex-associated DNA markers. About 1500bands were produced in total, nine primers were found yielding one or two DNA bands amplified in all nine male DNA bulks and absent in all female DNA bulks. These putatively male-associated markers were then scored in three different F1progenies, deriving from a cross between a common male parent and three different female plants. The sex of the progeny was accurately scored on the basis of the floral phenotype, and the presence of the nine male-associated markers was verified by RAPD analysis. In all three progenies examined, all the male plants showed the DNA markers previously identified by bulk segregant analysis (BSA) on the hemp varieties, while all the female plants lacked them. The fact that the association between these markers and the staminate phenotype is found when examining male plants of distantly related varieties, and that such linkage is never broken when different progenies are examined, strongly supports the hypothesis that the markers found are physically located on the Y chromosome, in a region excluded from recombination during meiosis. Another marker was shown to be present in the male parent, in all the male plants of each progeny, and in 50% of the female progenies, while it was absent in the female parent; the possible occurrence of markers deriving from multiple amplification sites of the genome is discussed. This revised version was published online in August 2006 with corrections to the Cover Date.  相似文献   

8.
以8077s与抗感的籼稻品种丰35亲本及杂交后自交所得的F2群体为材料，采用群分法（Bulked Segregant Analysis, BSA），从210个10mer随机引物，找到两个水稻苯达松敏感池和抗感池之间表现多态性的特异引物——S20和S316，分别产生的标记片段为S20-440和S316-590。它们与bel基因的连锁距离分别为12.132 cM和7.97 cM。对RAPD扩增标记的片段进行克隆、测序，根据测序结果合成两对特异性的SCAR引物，包含原有的RAPD序列。SC01引物在敏感单株中扩增出一条423 bp带；SC02引物在敏感单株中扩增出一条606 bp带，它们的SCAR标记与bel基因的连锁距离为10.66 cM和7.04 cM。应用SCAR标记对水稻恢复系进行了辅助选育。  相似文献   

9.
J. Hu    G. Li    D. Struss  C. F. Quiros 《Plant Breeding》1999,118(2):145-150
Breeding rapeseed for enhanced oil quality includes the development of varieties with low linolenic acid content. The breeder also aims to develop varieties with a high linoleic acid content because of its nutritional value. Restriction fragment length polymorphism (RFLP) and random amplified polymorphic DNA (RAPD) markers have been developed for linolenic acid content, but they are not best suited for a direct application in marker-assisted selection. The RFLP technique is too complex and time-consuming and RAPD markers lack codominance, precluding the distinction of homozygous from heterozygous individuals. In this report the conversion of a RAPD marker to a codominant sequence characterized amplified region (SCAR) marker named L1L9 is described. One of the alleles consisting of an 899 bp fragment (allele A), is associated with low linolenic acid content. The other allele consists of an 641 bp fragment (allele B) and is associated with high linolenic acid content. This marker explains approximately 25% of the genetic variation for this trait. Linkage analysis in the mapping population indicates that the SCAR marker probably tags an ω-3 desaturase gene in B. napus. Two RAPD markers were found to be associated with oleic/linoleic acid content. Markers M14-350 and I06-650 explained approximately 10% and 7% of the genetic variation for linoleic acid content, respectively. These two markers were found linked at 12.3cM in the segregating B. napus F2 progeny used for mapping. All the markers reported in this paper should be useful in breeding programmes for developing high linoleic and low linolenic acid rapeseed varieties.  相似文献   

10.
Sex type determination in papaya (Carica papaya L.) is very important for crop improvement processes because it accelerates the identification of the fruitful plants. The use of molecular technology provides a quick and reliable identification of sex types in plantlets growing in seedbeds. Random amplified polymorphic DNA (RAPD) markers were used to determine the sex types of Colombian cultivars of dioecious papaya genotypes. This species has three sex types (male, female and hermaphrodite) determined by a multiallelic locus. There are no morphological differences at the chromosome level; therefore the identification of sex types by chromosomal dimorphism is not possible. A RAPD marker of 900 bp was found in male plants, but not in females or hermaphrodites. From this RAPD marker a sequence characterized amplified region (SCAR) was developed and it was possible to amplify fragments from the genomes of male and hermaphrodite plants, but not the female ones. The results indicate that this new SCAR marker will be valuable to determine the sex type of papaya plants.  相似文献   

11.
Summary DNA polymorphism among five Asparagus officinalis L. cultivars-Imperial, Snow, Steline, UC-157 and Larac, as detected by random amplified polymorphic DNA (RAPD), is reported. Thirty one decamer primers were tested. and twenty six of them yielded amplification products. Fourteen primers gave products with at least one polymorphic DNA fragment. Among a total of 119 amplified fragments 33 were polymorphic. These RAPD markers enabled the identification of asparagus cultivars. Unique markers for cultivars were: Snow-bands 475 bp, 772 bp, 412 bp, 935 bp and 820 bp amplified by primers D5, OPA-07, OPA-09, OPA-10 and OPA-18, respectively. Steline-bands 645 bp, 680 bp and 997 bp amplified by primers A32, OPA-03 and OPA-09, respectively. A band 903 bp, amplitied by primer OPA-12, is a marker for Imperial, and a band 420 bp, amplified by primer D52, is a marker for Larac. Cultivar UC-157 could be identified by a combination of shared polymorphic bands. The pairwise marker difference between cultivars ranged from 0.08 to 0.17. A phenogram of the genetic relationship based on RAPD fits with the known origin of the cultivars.  相似文献   

12.
Aureobasidium pullulans strains Ach 1-1 and 1113-5 are two effective biocontrol agents against Botrytis cinerea and Penicillium expansum on stored apples. In the present work, a monitoring system allowing their identification and quantification was developed. The methodology used consisted of the development of both molecular markers and a semi-selective medium. The random amplified polymorphic DNA (RAPD) technique was applied to a collection of 15 strains of A. pullulans, including Ach 1-1 and 1113-5. Five specific RAPD fragments were amplified for strain Ach 1-1 and three others for strain 1113-5. Among them, a fragment of 528 bp specific to strain Ach 1-1 (generated with the OPR-13 RAPD primer) and another one of 431 bp specific to strain 1113-5 (amplified with the OPQ-03 RAPD primer) were selected, cloned, sequenced, and used to design sequence-characterized amplified region (SCAR) primers. Three different SCAR markers were amplified: two specific to strain Ach 1-1 (189 bp and 387 bp) and one specific to strain 1113-5 (431 bp). These SCAR primers can clearly identify strains Ach 1-1 and 1113-5 among 14 strains of A. pullulans and among eight yeast strains commonly present on apple fruit surfaces. Their selectivity was also tested using DNA extracted from epiphytic microflora of the apple surface. As a semi-selective medium, PDA medium supplemented with 0.5 mg L−1 euparen, 1 mg L−1 sumico, 2.5 mg L−1 hygromycin B, 30 mg L−1 streptomycin sulphate, and 1 mg L−1 cycloheximide was selected. It inhibited the development of the air microflora and appeared highly toxic for the epiphytic microflora of apple surface without altering the growth of the targeted strains Ach 1-1 and 1113-5. The combination of the semi-selective medium and SCAR markers provides a valuable monitoring tool to specifically identify and quantify A. pullulans strain Ach 1-1 and strain 1113-5 and could be used in future studies to evaluate their population dynamics under various laboratory and practical conditions.  相似文献   

13.
Similar to SCAR, an extended random primer amplified region (ERPAR) marker is a PCR amplified genomic DNA fragment at a single genetically defined locus. However, ERPAR uses specific primer pairs derived from RAPD primers by adding bases sequentially to their 3′-ends. As an example, an ERPAR marker was derived from a RAPD marker (OT11900) linked to a dominant male sterility gene in cabbage (Brassica oleracea var. capitata). After two cycles of base adding and primer pair screening, a primer pair (5′-TTCCCCGCGACT-3′and 5′-TTCCCCGCGAGA-3′) amplified a single intense band with the same size as OT11900. The identity of the new marker and OT11900 was verified by segregation analysis. The new marker amplified by this extended primer pair was named as EPT11900. The development of ERPAR exploits the importance of 3′-end bases of primers in PCR ERPAR shares advantages of SCAR, but eliminates the need for cloning and sequencing. It is a fast and universal way of converting RAPD markers into stable markers. This revised version was published online in July 2006 with corrections to the Cover Date.  相似文献   

14.
Evidence of gene introgression in apple using RAPD markers   总被引：4，自引：0，他引：4  
Summary A genomic remnant of Malus floribunda clone 821 introgressed into the cultivated apple M. x domestica Borkh. was identified using randomly amplified polymorphic DNA (RAPD) markers obtained by the polymerase chain reaction (PCR). Using a set of 59 oligonucleotide decamer primers, polymorphic DNA markers were identified among three pooled DNA samples. Based on the presence or absence of bands among bulked apple scab-resistant selections and cultivars, bulked scab-susceptible cultivars, and a M. floribunda clone 821 sample, one primer, A 15, identified amplified fragments in the scab-resistant bulked sample that was also unique to the M. floribunda clone 821. The unique band from M. floribunda clone 821 was amplified in four out of 17 scab-resistant selections/cultivars. This RAPD, designated OA15900, identifies an introgressed fragment that has as yet no known function.  相似文献   

15.
T. Markussen    J. Krüger    H. Schmidt  F. Dunemann 《Plant Breeding》1995,114(6):530-534
The availability of molecular markers linked to mildew resistance genes would enhance the efficiency of apple-breeding programmes. This investigation focuses on the identification of random amplified polymorphic DNA (RAPD) markers linked to the Pl1 gene for mildew resistance, which has introgressed from Malus robusta into cultivated apples. The RAPD marker technique was combined with a modified ‘bulked seg-regant analysis’ mapping strategy. About 850 random decamer primers used as single primers or in combinations were tested by PCR analysis on the basis of resistant and susceptible DNA pools. Selected primers producing RAPD fragments were applied in an additional selection step to M. robusta and genotypes representing intermediate breeding stages of the breeding population 93/9, for which a 1:1 segregation could be observed for the resistance trait. Seven RAPD markers, all representing introgressed DNA sequences from M. robusta, were identified and arranged with the Pl1 locus in a common linkage group. The two most tightly-linked RAPD markers, OPAT20450 and OPD21000 were mapped with a genetic distance of 4.5 and 5 cM, respectively, from the Pl1 gene. Both markers are suitable for marker-assisted selection in apple breeding. The polymorphic DNA fragment OPAT20450 was cloned and sequenced, and longer primers for the generation of a sequence-characterized amplified region (SCAR) marker have been constructed; this marker was easier to score than the original RAPD marker.  相似文献   

16.
干滟  曾凡亚  赵云  王茂林 《作物学报》2001,27(6):722-728
用100个随机引物对"蜀杂6号”父、母本进行RAPD扩增,选出5个能在父、母本中产生多态性扩增的引物.在杂种F1代中验证它们的特异性和稳定性,从中筛选出能从杂种F1代中稳定扩增"蜀杂6号”父、母本特异标记的随机引物GE204和GE222,并对它们扩增的父、母本特异标记片段进行克隆和测序.根据测序结果设计的特异序列扩增引物,将"蜀杂6  相似文献   

17.
Anthracnose, one of the destructive foliar diseases of sorghum growing in warm humid regions, is incited by the fungus Colletotrichum graminicola.The inheritance of anthracnose resistance was studied using the parental cultivars of Sorghum bicolor (L.) Moench, HC 136 (susceptible to anthracnose) and G 73 (anthracnose resistant). The F1 and F2 plants were inoculated with the local isolates of C. graminicola cultures. The F2 plants showed a segregation ratio of 3 (susceptible): 1(resistant) indicating that the locus for resistance to anthracnose in sorghum accession G 73 segregates as a recessive trait in a cross to susceptible cultivar HC 136. RAPD (random amplified polymorphic DNA) marker OPJ 011437 was identified as marker closely linked to anthracnose resistance gene in sorghum by bulked segregant analysis of HC 136 × G73 derived recombinant inbred lines (RILs) of sorghum. A total of 84 random decamer primers were used to screen polymorphism among the parental genotypes. Among these, only 24 primers were polymorphic. On bulked segregant analysis, primer OPJ 01 amplified a 1437 bp fragment only in resistant parent G 73 and resistant bulk. The marker OPJ 011437 was cloned and sequenced. The sequence of RAPD marker OPJ 011437 was used to generate specific markers called sequence characterized amplified regions (SCARs). A pair of SCAR markers SCJ 01-1 and SCJ 01-2 was developed using Mac Vector program. SCAR amplification of resistant and susceptible parents along with their respective bulks and RILs confirmed that SCAR marker SCJ 01 is at the same loci as that of RAPD marker OPJ 011437 and hence, is linked to anthracnose resistance gene. Resistant parent G 73 and resistant bulk amplified single specific band on PCR amplification using SCAR primer pairs. The RAPD marker OPJ 011437 was mapped at a distance of 3.26 cM apart from the locus governing anthracnose resistance on the sorghum genetic map by the segregation analysis of the RILs. Using BLAST program, it was found that the marker showed 100 per cent alignment with the contig{_}3966 located on the longer arm of chromosome 8 of sorghum genome. Therefore, these identified RAPD and SCAR markers can be used in the resistance-breeding program of sorghum anthracnose by marker-assisted selection.An erratum to this article can be found at  相似文献   

18.
利用RAPD技术, 从500个随机寡核苷酸引物(10聚体)中筛选出8个引物能在3个主栽的汕优系统杂交稻组合汕优63、 汕优64 和汕优晚3 及其亲本之间稳定地扩增出12个强的多态性标记。 利用这些多态性标记能够有效地区别汕优63、 汕优64 和汕优晚3 及其亲本。  相似文献   

19.
Novel male-specific molecular markers (MADC5, MADC6) in hemp   总被引：8，自引：0，他引：8  
Decamer RAPD primers were tested on dioecious and monoecious hemp cultivars to identify sex-specific molecular markers. Two primers (OPD05 and UBC354) generated specific bands in male plants. These two DNA fragments were isolated, cloned and sequenced. Both markers proved to be unique, since no sequence with significant homology to OPD05961 and UBC354151 markers were found in databases. These markers were named MADC3 (OPD05961) and MADC4 (UBC354151) (Male-Associated DNA from Cannabis sativa). The markers were converted into sequence-characterized amplified region (SCAR) markers. The SCAR markers correlated with the sex of the segregating F2 population and proved the tight linkage to the male phenotype. Results of F2 plant population analysis suggest these markers are to be linked to the Y chromosome. This revised version was published online in August 2006 with corrections to the Cover Date.  相似文献   

20.
In previous studies, we successfully transferred the P genome of Agropyron cristatum into wheat using wide hybridization methods. In the current investigation, repetitive sequences were cloned and DNA markers specific for the P genome of A. cristatum were developed. Three P genome-specific markers, designated OPX07-1036, OPX11-817 and OPC05-1539, were identified and isolated using random amplified polymorphic DNAs. The three markers were successfully amplified in all tested materials that contained Agropyron chromatin, such as Agropyron itself and wheat-Agropyron addition lines. These RAPD markers were converted into SCAR markers to be used in detection of P genome chromatin in wheat. In situ probing with fluorescent-labeled marker DNA has shown that they are distributed in all arms of Agropyron hence they will be useful in a variety of studies on introgressions of the P-genome chromatin into wheat.  相似文献   

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