| 03.09.2020 19:45 | | St Helens Town | | Cammell Laird 1907 | | 1:1, pen. 5:4 |
| St Helens Edge Green Street |
| 02.09.2020 20:00 | | Avro | | Bootle | | 1:2 |
| Oldham |
| 02.09.2020 20:00 | | Holbeach United | | Kirby Muxloe | | 1:0 |
| Holbeach |
| 02.09.2020 19:45 | | Crook Town AFC | | Yorkshire Amateur | | 1:1, pen. 5:4 |
| Crook |
| 02.09.2020 19:45 | | Guisborough Town | | Newton Aycliffe | | 1:0 |
| Guisborough |
| 02.09.2020 19:45 | | Stockton Town | | Shildon | | 3:0 |
| Stockton-on-Tees |
| 02.09.2020 19:45 | | Bishop Auckland | | Hall Road Rangers | | 1:0 |
| Bishop Auckland |
| 02.09.2020 19:45 | | Litherland REMYCA | | Ashton Athletic | | 3:0 |
| Litherland |
| 02.09.2020 19:45 | | Campion AFC | | Albion Sports | | 0:1 |
| Bradford |
| 02.09.2020 19:45 | | Carlton Town | | Loughborough University | | 1:1, pen. 4:2 |
| Carlton |
| 02.09.2020 19:45 | | Bottesford Town | | Selston | | 2:1 |
| Scunthorpe |
| 02.09.2020 19:45 | | Blackstones | | Boston Town | | 2:3 |
| Stamford |
| 02.09.2020 19:45 | | Pinchbeck United | | St Neots Town | | 0:4 |
| Pinchbeck |
| 02.09.2020 19:45 | | Godmanchester Rovers | | Ely City | | 1:1, pen. 2:4 |
| Godmanchester |
| 02.09.2020 19:45 | | Park View | | Hashtag United | | 1:2 |
| London |
| 02.09.2020 19:45 | | Baldock Town | | St Margaretsbury | | 2:2, pen. 2:4 |
| Hitchin |
| 02.09.2020 19:45 | | West Essex | | Crawley Green | | 6:3 |
| London |
| 02.09.2020 19:45 | | Ilford | | Halstead Town | | 1:3 |
| London |
| 02.09.2020 19:45 | | Roman Glass St George | | Flackwell Heath | | 1:2 |
| Bristol |
| 02.09.2020 19:45 | | Dunstable Town | | Thame Rangers | | 1:1, pen. 4:3 |
| Dunstable |
| 02.09.2020 19:45 | | Colliers Wood United | | Hollands & Blair | | 0:1 |
| Colliers Wood |
| 02.09.2020 19:45 | | Stansfeld | | Punjab United | | 3:3, pen. 4:3 |
| London |
| 02.09.2020 19:45 | | Guildford City | | Canterbury City | | 3:2 |
| Guildford |
| 02.09.2020 19:45 | | Balham | | Hassocks | | 2:2, pen. 3:5 |
| London |
| 02.09.2020 19:45 | | Baffins Milton Rovers | | AFC Stoneham | | 1:2 |
| Portsmouth |
| 02.09.2020 19:45 | | Westbury United | | Binfield | | 0:2 |
| Westbury |
| 02.09.2020 19:45 | | Sherborne Town | | Street | | 2:1 |
| Sherborne |
| 02.09.2020 19:30 | | Heaton Stannington | | West Auckland Town | | 2:3 |
| Newcastle upon Tyne |
| 02.09.2020 19:30 | | Mile Oak | | Beckenham Town | | 0:4 |
| Mile Oak |
| 02.09.2020 19:30 | | Tavistock | | Bradford Town | | 2:0 |
| Tavistock |
| 02.09.2020 19:30 | | Willand Rovers | | Bridport | | 2:1 |
| Willand |
| 01.09.2020 20:00 | | Thornaby | | Bridlington Town | | 2:0 |
| Thornaby-on-Tees |
| 01.09.2020 20:00 | | Skelmersdale United | | Penistone Church | | 3:2 |
| Skelmersdale |
| 01.09.2020 20:00 | | Kirkley & Pakefield | | Cogenhoe United | | 1:2 |
| Lowestoft |
| 01.09.2020 20:00 | | Portland United | | Clevedon Town | | 0:1 |
| Isle of Portland |
| 01.09.2020 19:45 | | Penrith | | Pickering Town | | 3:2 |
| Penrith |
| 01.09.2020 19:45 | | Northallerton Town | | Billingham Town | | 1:4 |
| Northallerton |
| 01.09.2020 19:45 | | Sunderland RCA | | Durham City | | 3:1 |
| Sunderland |
| 01.09.2020 19:45 | | Whickham | | West Allotment Celtic | | 0:0, pen. 2:4 |
| Whickham |
| 01.09.2020 19:45 | | Glasshoughton Welfare | | Knaresborough Town | | 1:1, pen. 2:4 |
| Glasshoughton |
| 01.09.2020 19:45 | | Hebburn Town | | Hemsworth Miners Welfare | | 4:1 |
| Hebburn |
| 01.09.2020 19:45 | | Consett AFC | | Ryhope Colliery Welfare | | 3:2 |
| Consett |
| 01.09.2020 19:45 | | Marske United | | North Shields | | 3:0 |
| Marske-by-the-Sea |
| 01.09.2020 19:45 | | Garforth Town | | Whitley Bay | | 0:3 |
| Garforth |
| 01.09.2020 19:45 | | Ashington | | Goole AFC | | 2:1 |
| Ashington |
| 01.09.2020 19:45 | | 1874 Northwich | | Warrington Rylands 1906 | | 1:6 |
| Winsford |
| 01.09.2020 19:45 | | Longridge Town | | Winsford United | | 2:1 |
| Longridge |
| 01.09.2020 19:45 | | Wythenshawe Amateurs | | Shelley Community | | 1:1, pen. 4:2 |
| Manchester |
| 01.09.2020 19:45 | | Runcorn Town | | Thackley | | 2:1 |
| Runcorn |
| 01.09.2020 19:45 | | Eccleshill United | | Silsden AFC | | 0:3 |
| Wrose |
| 01.09.2020 19:45 | | AFC Darwen | | Barnoldswick Town | | 2:4 |
| Darwen |
| 01.09.2020 19:45 | | Northwich Victoria | | Padiham | | 1:0 |
| Northwich |
| 01.09.2020 19:45 | | Daisy Hill | | Colne | | 0:2 |
| Westhoughton |
| 01.09.2020 19:45 | | Athersley Recreation | | Charnock Richard | | 1:4 |
| Athersley |
| 01.09.2020 19:45 | | Irlam | | Liversedge | | 2:1 |
| Salford |
| 01.09.2020 19:45 | | Maine Road | | Squires Gate | | 0:5 |
| Manchester |
| 01.09.2020 19:45 | | Congleton Town | | Burscough | | 1:1, pen. 3:2 |
| Congleton |
| 01.09.2020 19:45 | | Hanley Town | | Lye Town | | 3:1 |
| Hanley |
| 01.09.2020 19:45 | | Rugby Town | | Worcester City | | 0:2 |
| Rugby |
| 01.09.2020 19:45 | | Sporting Khalsa | | Boldmere St Michaels | | 2:1 |
| Willenhall |
| 01.09.2020 19:45 | | Highgate United | | AFC Bridgnorth | | 0:3 |
| Solihull |
| 01.09.2020 19:45 | | Stourport Swifts | | Coventry United | | 1:3 |
| Stourport-on-Severn |
| 01.09.2020 19:45 | | Gresley Rovers | | Wellington | | 2:0 |
| Church Gresley |
| 01.09.2020 19:45 | | Stone Old Alleynians | | Tividale | | 0:0, pen. 1:4 |
| Stone |
| 01.09.2020 19:45 | | Bewdley Town | | Black Country Rangers | | 1:5 |
| Bewdley |
| 01.09.2020 19:45 | | Romulus | | Coventry Sphinx | | 0:0, pen. 3:4 |
| Sutton Coldfield |
| 01.09.2020 19:45 | | Chelmsley Town | | Shifnal Town | | 0:3 |
| Coleshill |
| 01.09.2020 19:45 | | Westfields | | Brocton | | 2:0 |
| Hereford |
| 01.09.2020 19:45 | | Heather St John's | | Walsall Wood | | 0:1 |
| Heather |
| 01.09.2020 19:45 | | Whitchurch Alport | | Haughmond | | 3:1 |
| Whitchurch |
| 01.09.2020 19:45 | | Racing Club Warwick | | AFC Wulfrunians | | 3:0 |
| Warwick |
| 01.09.2020 19:45 | | Anstey Nomads | | Sleaford Town | | 3:0 |
| Anstey |
| 01.09.2020 19:45 | | Lutterworth Town | | Staveley Miners Welfare | | 2:0 |
| Lutterworth |
| 01.09.2020 19:45 | | Radford | | Shepshed Dynamo | | 1:1, pen. 3:4 |
| Nottingham |
| 01.09.2020 19:45 | | Spalding United | | Barton Town FC | | 1:3 |
| Spalding |
| 01.09.2020 19:45 | | Quorn | | Melton Town | | 2:0 |
| Quorn |
| 01.09.2020 19:45 | | AFC Mansfield | | Sherwood Colliery | | 2:1 |
| Clipstone The Forest Town Stadium |
| 01.09.2020 19:45 | | Long Eaton United | | Grimsby Borough | | 3:1 |
| Long Eaton |
| 01.09.2020 19:45 | | Newark | | Deeping Rangers | | 4:0 (1:0) |
| Newark Lowfields |
| 01.09.2020 19:45 | | Maltby Main | | Handsworth | | 2:1 |
| Maltby |
| 01.09.2020 19:45 | | West Bridgford | | Dunkirk | | 1:1, pen. 5:3 |
| West Bridgford Regatta Way Sports Ground |
| 01.09.2020 19:45 | | Leicester Nirvana | | Oadby Town | | 1:3 |
| Leicester |
| 01.09.2020 19:45 | | Long Melford | | Northampton ON Chenecks | | 3:2 |
| Long Melford |
| 01.09.2020 19:45 | | Whitton United | | Ipswich Wanderers | | 4:3 |
| Ipswich |
| 01.09.2020 19:45 | | Newmarket Town | | Walsham-le-Willows | | +:- |
| Newmarket |
| 01.09.2020 19:45 | | Hadleigh United | | Mildenhall Town | | 3:4 |
| Hadleigh |
| 01.09.2020 19:45 | | Peterborough Northern Star | | Potton United | | 0:1 |
| Peterborough |
| 01.09.2020 19:45 | | Diss Town | | Framlingham Town | | 1:0 |
| Diss |
| 01.09.2020 19:45 | | Thetford Town | | Wellingborough Town | | 2:0 |
| Blofield |
| 01.09.2020 19:45 | | Burton Park Wanderers | | AFC Sudbury | | -:+ |
| Burton Latimer |
| 01.09.2020 19:45 | | Gorleston | | Swaffham Town | | 5:0 |
| Gorleston |
| 01.09.2020 19:45 | | Haverhill Rovers | | Norwich United | | 2:1 |
| Haverhill |
| 01.09.2020 19:45 | | Eynesbury Rovers | | Desborough Town | | 7:3 |
| St Neots |
| 01.09.2020 19:45 | | Wellingborough Whitworth | | Harborough Town | | 1:6 |
| Wellingborough |
| 01.09.2020 19:45 | | Woodbridge Town | | Biggleswade United | | 2:2, pen. 6:7 |
| Woodbridge |
| 01.09.2020 19:45 | | Stowmarket Town | | Rothwell Corinthians | | 2:1 |
| Stowmarket |
| 01.09.2020 19:45 | | Wroxham | | Arlesey Town | | 5:0 |
| Wroxham |
| 01.09.2020 19:45 | | Redbridge | | Harpenden Town | | 0:2 |
| London |
| 01.09.2020 19:45 | | Harlow Town | | Enfield 1893 | | 5:1 |
| Harlow |
| 01.09.2020 19:45 | | Clapton | | Sporting Bengal United | | 2:2, pen. 5:6 |
| Forest Gate |
| 01.09.2020 19:45 | | Hoddesdon Town | | Clacton | | 1:2 |
| Hoddesdon |
| 01.09.2020 19:45 | | New Salamis | | Colney Heath | | 1:0 |
| London |
| 01.09.2020 19:45 | | Southend Manor | | Ware | | 0:5 |
| Southend-on-Sea |
| 01.09.2020 19:45 | | Walthamstow | | Maccabi London Lions | | 3:2 |
| London |
| 01.09.2020 19:45 | | Saffron Walden Town | | Little Oakley | | 4:2 |
| Saffron Walden |
| 01.09.2020 19:45 | | Stansted | | Takeley | | 3:2 |
| Stansted Mountfitchet |
| 01.09.2020 19:45 | | Cockfosters | | Stanway Rovers | | 4:2 |
| London |
| 01.09.2020 19:45 | | Hullbridge Sports | | Hadley | | 0:2 |
| Hullbridge |
| 01.09.2020 19:45 | | Sawbridgeworth Town | | Romford | | 2:1 |
| Sawbridgeworth |
| 01.09.2020 19:45 | | Brantham Athletic | | Benfleet | | 1:1, pen. 3:2 |
| Brantham |
| 01.09.2020 19:45 | | Oxhey Jets | | Bishop's Cleeve | | 1:0 |
| South Oxhey |
| 01.09.2020 19:45 | | Clanfield | | Long Crendon | | 0:1 |
| Clanfield |
| 01.09.2020 19:45 | | Leverstock Green | | Wembley | | 3:1 |
| Leverstock Green |
| 01.09.2020 19:45 | | Tuffley Rovers | | Harefield United | | 2:5 |
| Tuffley |
| 01.09.2020 19:45 | | Holmer Green | | Shrivenham | | 3:2 |
| Holmer Green |
| 01.09.2020 19:45 | | Winslow United | | Newport Pagnell Town | | 2:1 |
| Winslow |
| 01.09.2020 19:45 | | Cribbs | | Newent Town | | 3:1 |
| Bristol |
| 01.09.2020 19:45 | | Aylesbury Vale Dynamos | | Windsor | | 1:2 |
| Aylesbury |
| 01.09.2020 19:45 | | Lydney Town | | Fairford Town | | 0:1 |
| Lydney |
| 01.09.2020 19:45 | | Risborough Rangers | | Longlevens | | 4:1 |
| Princes Risborough |
| 01.09.2020 19:45 | | AFC Dunstable | | Hallen | | 4:0 |
| Dunstable |
| 01.09.2020 19:45 | | Royal Wootton Bassett Town | | Chipping Sodbury Town | | 2:2, pen. 4:1 |
| Wootton Bassett |
| 01.09.2020 19:45 | | Burnham | | Cheltenham Saracens | | 0:0, pen. 5:3 |
| Burnham |
| 01.09.2020 19:45 | | North Greenford United | | Brimscombe & Thrupp | | 1:2 |
| London |
| 01.09.2020 19:45 | | CB Hounslow United | | Banstead Athletic | | 6:1 |
| London |
| 01.09.2020 19:45 | | Billingshurst | | Westside | | 0:2 |
| Billingshurst |
| 01.09.2020 19:45 | | Lordswood | | Hanworth Villa | | 2:3 |
| Chatham |
| 01.09.2020 19:45 | | Molesey | | Southall | | 0:7 |
| Molesey |
| 01.09.2020 19:45 | | AFC Croydon Athletic | | Sutton Common Rovers | | 0:3 |
| London |
| 01.09.2020 19:45 | | Glebe | | Whyteleafe | | 1:3 |
| Bromley |
| 01.09.2020 19:45 | | Broadbridge Heath | | Raynes Park Vale | | 3:1 |
| Broadbridge Heath |
| 01.09.2020 19:45 | | Alfold | | Shoreham | | 9:1 |
| Alfold |
| 01.09.2020 19:45 | | Saltdean United | | Eastbourne Town | | 1:3 |
| Saltdean |
| 01.09.2020 19:45 | | Kennington | | Erith & Belvedere | | 0:1 |
| Ashford |
| 01.09.2020 19:45 | | Langney Wanderers | | Tower Hamlets | | 3:1 |
| Eastbourne |
| 01.09.2020 19:45 | | Abbey Rangers | | Welling Town | | 3:1 |
| Addlestone Addlestone Moor |
| 01.09.2020 19:45 | | Oakwood | | Cobham | | 0:2 |
| Crawley |
| 01.09.2020 19:45 | | Tunbridge Wells | | Erith Town | | 1:0 |
| Royal Tunbridge Wells |
| 01.09.2020 19:45 | | Newhaven | | Lingfield | | 1:1, pen. 4:2 |
| Newhaven |
| 01.09.2020 19:45 | | Broadfields United | | Loxwood | | 3:0 |
| London |
| 01.09.2020 19:45 | | Redhill | | Egham Town | | -:+ |
| Redhill |
| 01.09.2020 19:45 | | Fisher | | Horsham YMCA | | 2:0 |
| London |
| 01.09.2020 19:45 | | Corinthian | | Sheerwater | | 2:1 |
| Hartley Gay Dawn Farm |
| 01.09.2020 19:45 | | K Sports | | Steyning Town | | 0:2 |
| Aylesford |
| 01.09.2020 19:45 | | Eastbourne United Association | | Horley Town | | 0:1 |
| Eastbourne |
| 01.09.2020 19:45 | | Knaphill | | Deal Town | | 0:4 |
| Knaphill |
| 01.09.2020 19:45 | | Bearsted | | Peacehaven & Telscombe | | 3:5 |
| Otham |
| 01.09.2020 19:45 | | Spelthorne Sports | | Virginia Water | | 1:0 |
| Spelthorne |
| 01.09.2020 19:45 | | Sheppey United | | Sutton Athletic | | 1:0 |
| Isle of Sheppey |
| 01.09.2020 19:45 | | Corsham Town | | Farnham Town | | 2:2, pen. 4:3 |
| Corsham |
| 01.09.2020 19:45 | | Hamworthy United | | Calne Town | | 3:0 |
| Poole |
| 01.09.2020 19:45 | | Badshot Lea | | Amesbury Town | | 4:0 |
| Wrecclesham |
| 01.09.2020 19:45 | | Hamble Club | | Reading City | | 2:1 |
| Hamble-le-Rice |
| 01.09.2020 19:45 | | Frimley Green | | Fareham Town | | 1:0 |
| Frimley Green |
| 01.09.2020 19:45 | | Bashley | | Whitchurch United | | 8:2 |
| Bashley |
| 01.09.2020 19:45 | | Brockenhurst | | Lymington Town | | 0:1 |
| Brockenhurst |
| 01.09.2020 19:45 | | Basingstoke Town | | AFC Bournemouth | | 1:1, pen. 5:4 |
| Reading |
| 01.09.2020 19:45 | | Camberley Town | | Blackfield & Langley | | 0:0, pen. 5:4 |
| Camberley |
| 01.09.2020 19:45 | | Shaftesbury | | Fawley | | 2:2, pen. 5:6 |
| Shaftesbury |
| 01.09.2020 19:45 | | Tadley Calleva | | Alresford Town | | 1:0 |
| Tadley |
| 01.09.2020 19:45 | | Bemerton Heath Harlequins | | Horndean | | 2:1 |
| Bemerton |
| 01.09.2020 19:45 | | Christchurch | | Ascot United | | 1:1, pen. 7:6 |
| Christchurch |
| 01.09.2020 19:45 | | AFC Portchester | | Sandhurst Town | | 4:0 |
| Portchester |
| 01.09.2020 19:45 | | Keynsham Town | | Exmouth Town | | 1:3 |
| Keynsham |
| 01.09.2020 19:45 | | Odd Down | | Helston Athletic | | 1:4 |
| Bath |
| 01.09.2020 19:45 | | Wellington | | Bodmin Town | | 2:2, pen. 3:4 |
| Wellington |
| 01.09.2020 19:45 | | Plymouth Parkway | | Saltash United | | 1:1, pen. 4:5 |
| Plymouth |
| 01.09.2020 19:45 | | Millbrook | | Bridgwater Town | | 0:1 |
| Millbrook |
| 01.09.2020 19:45 | | Newton Abbot Spurs | | AFC St Austell | | 3:0 (2:0) |
| Newton Abbot |
| 01.09.2020 19:30 | | Newcastle Benfield | | Seaham Red Star | | 1:1, pen. 4:3 |
| Newcastle upon Tyne |
| 01.09.2020 19:30 | | Easington Sports | | Tring Athletic | | 2:0 |
| Easington |
| 01.09.2020 19:30 | | Ardley United | | Edgware Town | | 0:0 |
| Ardley |
| 01.09.2020 19:30 | | Leighton Town | | Abingdon United | | 3:0 |
| Barton-le-Clay |
| 01.09.2020 19:30 | | East Preston | | Chatham Town | | 0:2 |
| East Preston |
| 01.09.2020 19:30 | | Crowborough Athletic | | Crawley Down Gatwick | | 3:4 |
| Crowborough |
| 01.09.2020 19:30 | | AFC Uckfield Town | | Little Common | | 0:1 |
| Uckfield |
| 01.09.2020 19:30 | | Lancing | | Phoenix Sports | | 3:3, pen. 10:11 |
| Lancing |
| 01.09.2020 19:30 | | Pagham | | Fleet Town | | 2:1 |
| Pagham |
| 01.09.2020 19:30 | | Cowes Sports | | Totton & Eling | | 1:0 |
| Cowes |
| 01.09.2020 19:30 | | Buckland Athletic | | Bitton | | 1:1, pen. 3:5 |
| Newton Abbot |
| 01.09.2020 19:30 | | Wells City | | Bovey Tracey | | 2:0 |
| Wells |
| 01.09.2020 19:30 | | Shepton Mallet | | Torrington | | 3:0 |
| Shepton Mallet |
| 01.09.2020 19:30 | | Brislington | | Cadbury Heath | | 0:2 |
| Brislington |
| 31.08.2020 19:45 | | Woodford Town | | London Colney | | 3:1 |
| London Ashton Playing Fields |