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ARS Home » Southeast Area » Houma, Louisiana » Sugarcane Research » Research » Publications at this Location » Publication #340882

Research Project: Genetic Improvement of Sugarcane for Temperate Climates

Location: Sugarcane Research

Title: Whole genome amplification of single pollen grains from a sugarcane cultivar and analysis of the genetic relatedness based on SCoT markers

Author
item CHEN, PINGHUA - COLLABORATOR
item CHEN, RUKAI - COLLABORATOR
item XU, LIPING - COLLABORATOR
item WANG, HENGBO - COLLABORATOR
item CHEN, LIPING - COLLABORATOR
item LIN, BING - COLLABORATOR
item SHI, GUIJIAO - COLLABORATOR
item ZHANG, ZHUO - COLLABORATOR
item GAO, SANJI - COLLABORATOR
item GUO, JINLONG - COLLABORATOR
item Pan, Yong-Bao

Submitted to: Chinese Journal of Tropical Crops
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/31/2011
Publication Date: 12/31/2011
Citation: Chen, P., Chen, R., Xu, L., Wang, H., Chen, L., Lin, B., Shi, G., Zhang, Z., Gao, S., Guo, J., Pan, Y.-B. 2011. Whole genome amplification of single pollen grains from a sugarcane cultivar and analysis of the genetic relatedness based on SCoT markers. Chinese Journal of Tropical Crops. 32(11):2069-2075.

Interpretive Summary: Single pollens were suspended into a 30% sucrose solution from an anther of a sugacrane cultivar. A drop of the pollen suspension was spread into a glass slide. Pollens were collected individually under a dissecting microscope using a pair of special forceps and lysed in an alkali/detergent solution. The pollen lysate was then used as the template for Whole Genome Amplification. The results showed that the genomic DNA of 52 single pollens were successfully amplified. Amplified WGA DNA products of single pollen grains were verified and further analyzed with a special DNA marker called "start codon targeted polymorphism" (SCoT). Using the software NTSYSpc2.1, the genetic similarity coeficient values among single pollens were calculated. The values ranged from 0.0888 to 0.3654 with 36 single pollens clustered into two groups at the value of 0.187. The result indicated a high scale of single pollens shared the same genetic composition during reproductive cell division. The 52 single pollen grains were clustered into four groups and a probability distribution of the pollen grains was calculated based on the number of pollen grains in the four groups.

Technical Abstract: Single pollen grains were isolated from an intact anther of a sugarcane cultivar and collected using a pair of special forceps. The single pollen grains were lysed in an alkali/detergent solution respectively. The resulting solution was used as the template for Whole Genome Amplification. Genomic DNA of individual pollen grains was amplified successfully and WGA products of single pollen grains were tested by 5S rRNA-ITS primers to select those with high fidelity to the donor plant. The WGA products selected out were analyzed with SCoT markers and produced a mount of DNA polymorphism. Genetic similarity calculation was performed using the software NTSYSpc 2.1. The similarity coefficient values ranged from 0.0888~0.3654 and 69 percent of total pollen grains were clustered into two groups at the value of 0.187.The results indicated a high scale of pollen grains shared same gene loci during the meiotic segregation stage. The 52 single pollen grain samples were clustered into four groups based on the homology tree. A probability distribution of the pollen grains was calculated based on the number of pollen grains in the four groups.