Nghiên cứu khu hệ vi sinh vật vùng rễ cây nghệ vàng curcuma longa l nhằm định hướng tăng năng suất và chất lượng củ nghệ

BỘ GIÁO DỤC VÀ ĐÀO TẠO VIỆN HÀN LÂM KHOA HỌC VÀ CÔNG NGHỆ VIỆT NAM HỌC VIỆN KHOA HỌC VÀ CÔNG NGHỆ - Hoàng Kim Chi NGHIÊN CỨU KHU HỆ VI SINH VẬT VÙNG RỄ CÂY NGHỆ VÀNG Curcuma longa L NHẰM ĐỊNH HƯỚNG TĂNG NĂNG SUẤT VÀ CHẤT LƯỢNG CỦ NGHỆ LUẬN ÁN TIẾN SĨ SINH HỌC Hà Nội – 2020 BỘ GIÁO DỤC VÀ ĐÀO TẠO VIỆN HÀN LÂM KHOA HỌC VÀ CÔNG NGHỆ VIỆT NAM HỌC VIỆN KHOA HỌC VÀ CÔNG NGHỆ Hoàng Kim Chi NGHIÊN CỨU KHU HỆ VI SINH VẬT VÙNG RỄ CÂY NGHỆ VÀNG Curcuma longa L NHẰM ĐỊNH HƯỚNG TĂNG NĂNG SUẤT VÀ CHẤT LƯỢNG CỦ NGHỆ Chuyên ngành: Vi sinh vật học Mã số: 42 01 07 LUẬN ÁN TIẾN SĨ SINH HỌC NGƯỜI HƯỚNG DẪN KHOA HỌC: GS TS Lê Mai Hương TS Trần Thị Như Hằng Hà Nội – 2020 MỤC LỤC Trang MỤC LỤC .i LỜI CAM ĐOAN v LỜI CẢM ƠN .vi DANH MỤC CÁC KÝ HIỆU VÀ CHỮ VIẾT TẮT vii DANH MỤC CÁC BẢNG ix DANH MỤC CÁC HÌNH VẼ, ĐỒ THỊ xi MỞ ĐẦU CHƯƠNG TỔNG QUAN 1.1 Một số vấn đề liên quan đến phân bón suất trồng 1.1.1 Phân bón hóa học, phân bón hữu hay phân bón sinh học? .3 1.1.2 Quản lý dinh dưỡng tổng hợp 1.1.3 Một số nhóm VSV hữu hiệu sản xuất phân sinh học 1.2 Cây nghệ vàng Cucuma longa L hợp chất curcumin củ nghệ 1.2.1 Hợp chất curcumin củ nghệ vàng 10 1.2.1.1 Thành phần hóa học củ nghệ vàng 10 1.2.1.2 Hoạt tính sinh y dược học hoạt chất curcumin củ nghệ vàng .11 1.2.2 Một số khía cạnh liên quan đến suất nghệ vàng 12 1.2.2.1 Khía cạnh sinh học 12 1.2.2.2 Khía cạnh nơng nghiệp 14 1.2.2.3 Chất lượng đất trồng, phân bón suất nghệ vàng 15 1.3 Vi sinh vật vùng rễ cộng sinh với nghệ vàng 16 1.3.1 Vi sinh vật vùng rễ thực vật .16 1.3.2 Một số nhóm VSVVR liên quan đến suất nghệ vàng .19 1.3.2.1 Vi khuẩn rễ kích thích sinh trưởng nghệ vàng 20 1.3.2.2 Nấm rễ cộng sinh nghệ vàng 21 1.3.3 Mối liên hệ khu hệ vi sinh vật cộng sinh hàm lượng curcumin nghệ vàng 22 1.3.3.1 Sự tổng hợp curcuminoid nghệ vàng 22 1.3.3.2 Vi sinh vật tạo thành curcumin nghệ 23 1.4 Bước phát triển công nghệ nghiên cứu khu hệ VSV vùng rễ 24 1.4.1 Một số phương pháp nghiên cứu khu hệ VSV đất 24 1.4.2 Metagenomics nghiên cứu khu hệ VSV vùng rễ 25 i CHƯƠNG VẬT LIỆU VÀ PHƯƠNG PHÁP NGHIÊN CỨU 2.1Nguyên vật liệu 2.1.1 Giống 2.1.1.1 Đặc điểm sinh học 2.1.1.2 Mùa vụ chế độ chăm sóc Hóa ch 2.1.2 2.2Phương pháp nghiên cứu Thu mẫ 2.2.1 Thu mẫ 2.2.1.1 2.2.1.2 Xác định số lý hóa đất • Thành • pH • Hàm lư • Hàm lư • Hàm lư • Hàm lư • Hàm lư • Hàm lư 2.2.2 Nghiên 2.2.2.2 suất Bố tr 32 2.2.2.3 Xác định tiêu nghiên cứu • Tiến hà • Xác địn • Xác địn 2.2.3 VSVVR nghệ Nghiên 2.2.3.1 Tách DNA tổng số từ mẫu đất vùng rễ nghệ 2.2.3.2 Giải trình tự metagenome amplicons 2.2.3.3 Phân tích tin sinh học • Tiền xử • Xác địn • Phân tí Phân lậ 2.2.4 2.2.4.1 Phân lập vi khuẩn vùng rễ có khả hịa tan phosphate vơ ii 2.2.4.2 Xác định khả sinh IAA kháng nấm gây bệnh thực vật 2.2.4.3 Xác định đặc tính sinh lý, sinh hóa PGPR 2.2.4.4 Xác định tên phân loại dựa trình tự đoạn gene 16S rDNA Phân lập nấm AM 2.2.5 2.2.5.1 Phân lập bào tử nấm AM từ mẫu đất vùng rễ nghệ 2.2.5.2 Khảo sát đa dạng nấm AM in situ dựa trình tự vùng gen 18S rRNA 2.2.5.3 Xác định tên phân loại nấm AM kỹ thuật Nested PCR 2.2.5.4 Xác định đặc điểm hình thái bào tử nấm AM Nghiên cứu tạo 2.2.6 2.2.6.1 Tạo chế phẩm VSV vùng rễ cho nghệ • Xác định khả • Nhân ni nấm AM • Nhân sinh khối, tạo • Thu hồi bào tử, tạo • Phối trộn, tạo chế p 2.2.6.2 Thử nghiệm hiệu chế phẩm VSV vùng rễ cho nghệ • Bố trí thí nghiệm • Xác định tiê • Xác định khả CHƯƠNG KẾT QUẢ VÀ THẢO LUẬN 3.1 Khảo sát ảnh hưởng chế độ bón phân đạm hóa học đến suất nghệ 47 3.1.1 Xác định chất lượng nghệ 3.1.2 Điều kiện thổ nhưỡng khu 3.1.2.1 Đặc điểm khí hậu 3.1.2.2 Khảo sát số môi trường đất trồng nghệ 3.1.3 Xác định suất nghệ t 3.1.4 Xác định suất curcu 3.2Khảo sát số nhóm vi sinh vật hữu hiệu vùng rễ nghệ 3.2.1 Khảo sát vi khuẩn rễ kích 54 3.2.1.1 Phân lập tuyển chọn vi khuẩn PGPR 3.2.1.2 Đặc điểm hình thái, sinh lý, sinh hóa phân loại học chủng vi khuẩn lựa chọn 57 3.2.2 Khảo sát nấm AM vùng rễ nghệ 62 iii 3.2.2.1 Phân lập khả 3.2.2.2 Đặc điểm hình t vùng rễ nghệ 3.3Nghiên cứu tác động chế độ bón phân N đến đa dạng khu hệ vi sinh vật vùng rễ nghệ 3.3.1 Kết tách DNA 3.3.2 Kết giải trình t 3.3.3 Đánh giá đa dạng bón phân N 3.3.3.1 Phân tích 3.3.3.2 Thành phần phân 3.3.4 phân N 77 Đánh giá đa dạng 3.3.4.1 Phân tích 3.3.4.2 Thành phần phân 3.3.5 Luận giải tác độ vùng rễ nghệ 3.3.5.1 Chế độ bón phân 3.3.5.2 Cơ chế tác động c 3.3.6 nghệ Khác biệt 3.3.6.1 Vi khuẩn thuộc họ 3.3.6.2 Nấm thuộc ngành 3.4 Bước đầu nghiên cứu tạo chế phẩm sinh học làm tăng suất nghệ 87 3.4.1 Thử nghiệm tính an t 3.4.2 Tạo chế phẩm sinh họ 3.4.3 Thử nghiệm tác động KẾT LUẬN VÀ KIẾN NGHỊ DANH MỤC CÁC CÔNG TRÌNH LIÊN QUAN ĐẾN LUẬN ÁN TÀI LIỆU THAM KHẢO iv LỜI CAM ĐOAN Tơi xin cam đoan: Đây cơng trình nghiên cứu riêng hướng dẫn khoa học GS TS Lê Mai Hương TS Trần Thị Như Hằng Các số liệu kết thu luận án hoàn toàn trung thực chưa cơng bố cơng trình khác Tác giả luận án Hoàng Kim Chi v LỜI CẢM ƠN Luận án hoàn thành Học viện Khoa học Công nghệ – Viện Hàn lâm Khoa học Công nghệ Việt Nam Với kính trọng, lịng biết ơn chân thành sâu sắc nhất, tơi xin bày tỏ lịng biết ơn tới GS TS Lê Mai Hương TS Trần Thị Như Hằng người thầy hướng dẫn tận tình tạo điều kiện giúp đỡ thời gian thực luận án Tôi xin trân trọng cảm ơn quan tâm giúp đỡ Ban lãnh đạo Học viện Khoa học Cơng nghệ, Viện Hóa học hợp chất thiên nhiên Viện Công nghệ Sinh học tạo điều kiện giúp đỡ tơi hồn thành luận án Tơi xin bày tỏ lịng biết ơn tới tập thể cán Phòng Sinh học thực nghiệm, Viện Hóa học hợp chất thiên nhiên giúp đỡ tơi nhiệt tình suốt thời gian thực luận án Tôi xin gửi lời cảm ơn đặc biệt tới TS Eiko Kuramae, TS Trần Hồ Quang, TS Phạm Hồng Nam, TS Hồ Bích Hải, TS Tjalf de Boer, GS Bram Brouwer cho giúp đỡ trình tơi thực luận án Tơi xin cảm ơn đề tài nghiên cứu mã số ĐTĐLCN.14/14 Bộ Khoa học Công nghệ, đề tài VAST 0205/17-18 Viện Hàn lâm Khoa học Công nghệ, đề tài “Nghiên cứu sản xuất ứng dụng chế phẩm vi sinh vật phân hủy phốt hữu (OP) góp phần giảm thiểu nhiễm mơi trường tăng suất trồng” Bộ Nông nghiệp Phát triển Nông thôn tài trợ cho nghiên cứu luận án Tôi xin chân thành cảm ơn đồng nghiệp, bạn bè gần xa cổ vũ, động viên tơi hồn thành tốt luận án Cuối cùng, tơi xin bày tỏ lịng kính trọng biết ơn sâu sắc đến gia đình tơi, người khơng ngại khó khăn vất vả, ln giúp đỡ tạo điều kiện cho thực tốt luận án Tôi xin trân trọng cảm ơn! 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DỤC VÀ ĐÀO TẠO VI? ??N HÀN L? ?M KHOA HỌC VÀ CÔNG NGHỆ VI? ??T NAM HỌC VI? ??N KHOA HỌC VÀ CÔNG NGHỆ Hoàng Kim Chi NGHIÊN CỨU KHU HỆ VI SINH VẬT VÙNG RỄ CÂY NGHỆ VÀNG Curcuma longa L NHẰM ĐỊNH... cứu mối liên hệ chế độ bón phân đạm hóa học suất nghệ; (ii) Phân l? ??p nghiên cứu hoạt tính số nhóm vi sinh vật vùng rễ nghệ vàng; (iii) Nghiên cứu đa dạng khu hệ vi sinh vật vùng rễ mối liên hệ. .. công nghệ sinh học để l? ?m tăng phẩm chất hàm l? ?ợng chất đối tượng nghiên cứu 1.2.1 Hợp chất curcumin củ nghệ vàng 1.2.1.1 Thành phần hóa học củ nghệ vàng Thành phần hóa học củ nghệ vàng C longa