Complete Chloroplast Genome Sequence of Erigeron breviscapus and Characterization of Chloroplast Regulatory Elements

Erigeron breviscapus is a famous medicinal plant. However, the limited chloroplast genome information of E. breviscapus, especially for the chloroplast DNA sequence resources, has hindered the study of E. breviscapus chloroplast genome transformation. Here, the complete chloroplast (cp) genome of E....

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Autores principales:	Yifan Yu, Zhen Ouyang, Juan Guo, Wen Zeng, Yujun Zhao, Luqi Huang
Formato:	article
Lenguaje:	EN
Publicado:	Frontiers Media S.A. 2021
Materias:	Erigeron breviscapus chloroplast genome sequencing chloroplast regulatory elements gene assembly prokaryotic expression Plant culture SB1-1110
Acceso en línea:	https://doaj.org/article/7ca4a1a15a4c4e97827b8ea7a1f2aba4
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spelling	oai:doaj.org-article:7ca4a1a15a4c4e97827b8ea7a1f2aba42021-12-01T03:36:31ZComplete Chloroplast Genome Sequence of Erigeron breviscapus and Characterization of Chloroplast Regulatory Elements1664-462X10.3389/fpls.2021.758290https://doaj.org/article/7ca4a1a15a4c4e97827b8ea7a1f2aba42021-11-01T00:00:00Zhttps://www.frontiersin.org/articles/10.3389/fpls.2021.758290/fullhttps://doaj.org/toc/1664-462XErigeron breviscapus is a famous medicinal plant. However, the limited chloroplast genome information of E. breviscapus, especially for the chloroplast DNA sequence resources, has hindered the study of E. breviscapus chloroplast genome transformation. Here, the complete chloroplast (cp) genome of E. breviscapus was reported. This genome was 152,164bp in length, included 37.2% GC content and was structurally arranged into two 24,699bp inverted repeats (IRs) and two single-copy areas. The sizes of the large single-copy region and the small single-copy region were 84,657 and 18,109bp, respectively. The E. breviscapus cp genome consisted of 127 coding genes, including 83 protein coding genes, 36 transfer RNA (tRNA) genes, and eight ribosomal RNA (rRNA) genes. For those genes, 95 genes were single copy genes and 16 genes were duplicated in two inverted regions with seven tRNAs, four rRNAs, and five protein coding genes. Then, genomic DNA of E. breviscapus was used as a template, and the endogenous 5' and 3' flanking sequences of the trnI gene and trnA gene were selected as homologous recombinant fragments in vector construction and cloned through PCR. The endogenous 5' flanking sequences of the psbA gene and rrn16S gene, the endogenous 3' flanking sequences of the psbA gene, rbcL gene, and rps16 gene and one sequence element from the psbN-psbH chloroplast operon were cloned, and certain chloroplast regulatory elements were identified. Two homologous recombination fragments and all of these elements were constructed into the cloning vector pBluescript SK (+) to yield a series of chloroplast expression vectors, which harbored the reporter gene EGFP and the selectable marker aadA gene. After identification, the chloroplast expression vectors were transformed into Escherichia coli and the function of predicted regulatory elements was confirmed by a spectinomycin resistance test and fluorescence intensity measurement. The results indicated that aadA gene and EGFP gene were efficiently expressed under the regulation of predicted regulatory elements and the chloroplast expression vector had been successfully constructed, thereby providing a solid foundation for establishing subsequent E. breviscapus chloroplast transformation system and genetic improvement of E. breviscapus.Yifan YuYifan YuZhen OuyangJuan GuoWen ZengYujun ZhaoLuqi HuangLuqi HuangFrontiers Media S.A.articleErigeron breviscapuschloroplast genome sequencingchloroplast regulatory elementsgene assemblyprokaryotic expressionPlant cultureSB1-1110ENFrontiers in Plant Science, Vol 12 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Erigeron breviscapus chloroplast genome sequencing chloroplast regulatory elements gene assembly prokaryotic expression Plant culture SB1-1110
spellingShingle	Erigeron breviscapus chloroplast genome sequencing chloroplast regulatory elements gene assembly prokaryotic expression Plant culture SB1-1110 Yifan Yu Yifan Yu Zhen Ouyang Juan Guo Wen Zeng Yujun Zhao Luqi Huang Luqi Huang Complete Chloroplast Genome Sequence of Erigeron breviscapus and Characterization of Chloroplast Regulatory Elements
description	Erigeron breviscapus is a famous medicinal plant. However, the limited chloroplast genome information of E. breviscapus, especially for the chloroplast DNA sequence resources, has hindered the study of E. breviscapus chloroplast genome transformation. Here, the complete chloroplast (cp) genome of E. breviscapus was reported. This genome was 152,164bp in length, included 37.2% GC content and was structurally arranged into two 24,699bp inverted repeats (IRs) and two single-copy areas. The sizes of the large single-copy region and the small single-copy region were 84,657 and 18,109bp, respectively. The E. breviscapus cp genome consisted of 127 coding genes, including 83 protein coding genes, 36 transfer RNA (tRNA) genes, and eight ribosomal RNA (rRNA) genes. For those genes, 95 genes were single copy genes and 16 genes were duplicated in two inverted regions with seven tRNAs, four rRNAs, and five protein coding genes. Then, genomic DNA of E. breviscapus was used as a template, and the endogenous 5' and 3' flanking sequences of the trnI gene and trnA gene were selected as homologous recombinant fragments in vector construction and cloned through PCR. The endogenous 5' flanking sequences of the psbA gene and rrn16S gene, the endogenous 3' flanking sequences of the psbA gene, rbcL gene, and rps16 gene and one sequence element from the psbN-psbH chloroplast operon were cloned, and certain chloroplast regulatory elements were identified. Two homologous recombination fragments and all of these elements were constructed into the cloning vector pBluescript SK (+) to yield a series of chloroplast expression vectors, which harbored the reporter gene EGFP and the selectable marker aadA gene. After identification, the chloroplast expression vectors were transformed into Escherichia coli and the function of predicted regulatory elements was confirmed by a spectinomycin resistance test and fluorescence intensity measurement. The results indicated that aadA gene and EGFP gene were efficiently expressed under the regulation of predicted regulatory elements and the chloroplast expression vector had been successfully constructed, thereby providing a solid foundation for establishing subsequent E. breviscapus chloroplast transformation system and genetic improvement of E. breviscapus.
format	article
author	Yifan Yu Yifan Yu Zhen Ouyang Juan Guo Wen Zeng Yujun Zhao Luqi Huang Luqi Huang
author_facet	Yifan Yu Yifan Yu Zhen Ouyang Juan Guo Wen Zeng Yujun Zhao Luqi Huang Luqi Huang
author_sort	Yifan Yu
title	Complete Chloroplast Genome Sequence of Erigeron breviscapus and Characterization of Chloroplast Regulatory Elements
title_short	Complete Chloroplast Genome Sequence of Erigeron breviscapus and Characterization of Chloroplast Regulatory Elements
title_full	Complete Chloroplast Genome Sequence of Erigeron breviscapus and Characterization of Chloroplast Regulatory Elements
title_fullStr	Complete Chloroplast Genome Sequence of Erigeron breviscapus and Characterization of Chloroplast Regulatory Elements
title_full_unstemmed	Complete Chloroplast Genome Sequence of Erigeron breviscapus and Characterization of Chloroplast Regulatory Elements
title_sort	complete chloroplast genome sequence of erigeron breviscapus and characterization of chloroplast regulatory elements
publisher	Frontiers Media S.A.
publishDate	2021
url	https://doaj.org/article/7ca4a1a15a4c4e97827b8ea7a1f2aba4
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Complete Chloroplast Genome Sequence of Erigeron breviscapus and Characterization of Chloroplast Regulatory Elements

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