2018. Jihyun Park, Seonghyun Lee, Nabin Won, Eunji Shin, Soo-Hyun Kim, Min-Young Chun, Jungyeun Gu, Gun-Young Jung, Kwang-Il Lim * and Kyubong Jo *. "Single-molecule DNA visualization using AT-specific red and non-specific green DNA-binding fluorescent proteins" / Analyst (Top 15% journal)

October 7, 2018

 

 

The recent advances in the single cell genome analysis are generating a considerable amount of novel insights into complex biological systems. However, there are still technical challenges because each cell has a single copy of DNA to be amplified in most single cell genome analytical methods. In this paper, we present a novel approach to directly visualize a genomic map on a large DNA molecule instantly stained with red and green DNA-binding fluorescent proteins without DNA amplification. For this visualization, we constructed a few types of fluorescent protein-fused DNA-binding proteins: H-NS (histone-like nucleoidstructuring protein), DNA-binding domain of BRCA1 (breast cancer 1), high mobility group-1 (HMG), and lysine tryptophan (KW) repeat motif. Because H-NS and HMG preferentially bind A/T-rich regions, we combined A/T specific binder (H-NS-mCherry and HMG-mCherry as red color) and a non-specific complementary DNA binder (BRCA1-eGFP and 2(KW)2-eGFP repeat as green color) to produce a sequence-specific two-color DNA physical map for efficient optical identification of single DNA molecules.

 

 

 

DOI: 10.1039/C8AN01426D

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