Publication

Single-molecule DNA visualization using AT-specific red and non-specific green DNA-binding fluorescent proteins

조회수 : 75 등록일 : 2019.04.15 00:00

저자 : Park, J (Park, Jihyun); Jo, K (Jo, Kyubong)
출처 : ANALYST
출판일 : 2019.02.07


Single-molecule DNA visualization using AT-specific red and non-specific green DNA-binding fluorescent proteins


Park, J (Park, Jihyun)1,2 ] Lee, S (Lee, Seonghyun)1,2 ] Won, N (Won, Nabin)1,2 ] Shin, E (Shin, Eunji)1,2 ] Kim, SH (Kim, Soo-Hyun)3 ] Chun, MY(Chun, Min-Young)1,2 ] Gu, J (Gu, Jungyeun)1,2 ] Jung, GY (Jung, Gun-Young)4 ] Lim, KI (Lim, Kwang-Il)3 ] Jo, K (Jo, Kyubong)1,2 ]


[ 1 ] Sogang Univ, Dept Chem, 1 Shinsudong, Seoul 04107, South Korea
[ 2 ] Sogang Univ, Interdisciplinary Program Integrated Biotech, 1 Shinsudong, Seoul 04107, South Korea
[ 3 ] Sookmyung Womens Univ, Dept Chem & Biol Engn, Cheongpa Ro 47,Gil 100, Seoul 04310, South Korea
[ 4 ] GIST, Sch Mat Sci & Engn, Gwangju 61005, South Korea



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 nucleoid-structuring 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.






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