43 6 Vol143,No16 2006 11 ACTA PEDOLOGICA SINICA Nov.,2006 3 g (, 210095) CONSTRUCTION OF TYPICAL RED SOIL2DERIVED METAGENOMIC L IBRARY AND FUNCTIONAL ANALYSIS IN JIANGXI PROVINCE Huang Tingting Cui Zhongli Zhang Lu Cao Hui g Li Shunpeng ( Key Laboratory of Microbiological Engineering of Agricultural Environment, Ministry of Agriculture, Nanjing Agricultural University, Nanjing 210095, China) ;DNA ; ; S15413 ; Q93811 A, 4 10 30 6 10 30, 10 6 10 8 [1 ], [2 ], (metagenomics),, [3,4 ], 42, [5 9 ] [10,11 ], DNA, TA,, 1 111,, 1718, 1 785 mm, 4 6, 3510 5416 m, 5 8 [12 ], ph 4141, 1132 g kg - 1, 0167 g kg - 1, 20199 g kg - 1, 0 5 cm 5 15 cm 15 30 cm( a, b, c),,, 4 mm, - 70 DNA 1 3 (40371069) ( KZCX32SW2427) (2003033495) g,e2mail : hcao @njau. edu. cn : (1982 ),,,, :2005-12 - 01 ; :2006-08 - 09
1038 43 1 ( :cfu g - 1 ) (cm) a 0 5 812 10 7 213 10 7 1134 10 6 b 5 15 4175 10 7 919 10 6 2145 10 5 c 15 30 1155 10 7 2165 10 6 215 10 4 :a,b,c a 0 5 cm, b 5 15 cm, c 15 30 cm 112 puc 18 (Amp + ) DH5 ; Taq TA pmd182t Vector Takara, (Amp) CTAB ( ) SDS( ) Tris ( ) EDTA ; K Merk ; 113 DNA DNA Blending method [13 ] 260 nm DNA, 230 nm 280 nm DNA 16S rdna DNA DNA, [14 ] 114 11411 1 11412 BamHI DNA, DNA 2 6 kb DNA 45 min, Taq da : 10 Taq 5 l, Mg 2 + (25 mmol L - 1 ) 4 l, dntp (25 mmol L - 1 ) 5 l, DNA 10 l, Taq (5 U l - 1 ) 015 l, 50 l :94 3 min ;72 45 min ;10 10 min 11413 Inoue DH5 [14 ] ; T (pmd218,takara) 16, DH5,37 12 14 h ; LB,, 30 % - 70 115 DNA ;,, BLAST BioEdit ClustalX (ORF) 116 GenBank, GeneBank H3, H5, H10, H222, H223, H227, X6, Q41, (Accession number) : DQ185049, DQ185050, DQ185051, DQ185052, DQ185053,DQ195083,DQ195084,DQ195085 2 1 211 DNA 2 DNA 23 kb, DNA 6185 0124 g, DNA PCR
6 : 1039, DNA [15,16 ] A 260 / A 230 A 260 / A 280 DNA [17 ] DNA ( 2), 16S rdna, DNA 0 5 cm, 5 15 cm DNA 2 DNA a,b,c : DNA ;M DNA/ Hind Marker 2 DNA A 260 / A 230 A 260 / A 280 DNA g g - 1 a 11065 11001 7156 b 11062 11433 6185 c 11015 11374 1122 212, DNA, TA,, DNA DNA( 2 DNA ), DNA,,, DNA TA [18 ], BamH DNA, 2 6 kb, Taq PCR da, da, T, 1g 15 000, 80 % ( 4), DNA DNA, DNA G + C %, DNA ( Pst, EcoR, BamH Hind ) DNA, BamH ( 3) 3 DNA 1 : Pst ;2 : EcoR ;3 : Hind ;4 : BamH ; 5 : DNA ;6 : DNA/ Hind Marker 4 1,21 : DNA/ Hind Marker ;2 10, 13 20 : 16 ; 11,12 : puc18 213 10,, BLAST 10, 2 DNA, 8, BioEdit ClustalX (Opening reading frame,
1040 43 ORF), ( 3) 3 ORF ORF 1) S 2) E 3) H222 DQ185052 Frame 1, 239aa 3311 814 Frame21, 98aa 172 2 10-42 Frame22, 140aa IcmF, 120 1 10-26 H223 DQ185053 Frame 1, 117aa 106 2 10-22 H227 DQ195083 Frame 1, 169aa WD40 8617 3 10-16 Frame21, 74aa 3217 312 Frame22, 129aa DNA I 3311 214 H3 DQ185049 Frame 1, 87aa, PDZ C 3311 218 Frame21, 87aa ATP DNA 7917 3 10-14 H5 DQ185050 Frame 1, 74aa LacOPZ2 5913 4 10-8 H10 DQ185051 Frame 1, 131aa 3413 111 Frame21, 131aa 2 67 2 10-10 X6 DQ195084 3 3 Q41 DQ195085 3 3 Frame 3, 68 aa ATP cassette,sub2family B (MDR/ TAP) 3213 411 1) ORF, frame 1, 2 ORF, frame21, - 2 ORF 3 3 ORF 2) S (bits score),, 3) E ( E value),e ; S E 3,, DNA, H2-3, 29 % 55 % 9 ( 5),,, (S) ( H) (D),, 5 H223 9 1 : H223 ;2 : Bradyrhizobium japonicum USDA 110 ;3 : Magnetospirillum magnetotacticum ;4 : Paracoccus denitrificans ;5 : Rhodospirillum rubrum ;6 : Rhizobium sp ; 7 : Neisseria meningitidis Z2491 ;8 : Neisseria gonorrhoeae FA 1090 ;9 : Thiobacillus denitrificans ATCC 25259 ;10 : Mesorhizobium sp1 BNC1 H222 ORF ( ) (
6 : 1041 ) ( 6),,, ( Acidobacteria group) ( Solibacteres) 81 %, 40 % 50 % 16S rdna,, 6 H222 ORF 1 : Streptomyces coelicolor A3(2) ;2 : Streptomyces avermitilis MA24680 ;3 : Nocardioides sp1 JS614 ;4 : Bifidobacterium longum DJO10A ;5 :Lactobacillus casei ATCC 334 ;6 : Desulfitobacterium hafniense DCB22 ;7 : Bacillus halodurans C2125 ;8 :H222 ;9 : Solibacter usitatus Ellin6076 ;10 : Thiobacillus denitrificans ATCC 25259 3 DNA ( ) ( ) DNA DNA,, PCR ; DNA DNA,, [13 ] DNA, DNA,, DNA 10 kb,,, PCR da dt,, T,,,, [19 ],,,,,,
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